ehec o157 h7 edl933 strain atcc 700927  (ATCC)


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    ATCC ehec o157 h7 edl933 strain atcc 700927
    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC <t>EDL933</t> (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Ehec O157 H7 Edl933 Strain Atcc 700927, supplied by ATCC, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli"

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-12-231

    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC EDL933 (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Figure Legend Snippet: SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC EDL933 (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Techniques Used: Expressing, Mutagenesis, Labeling

    Increased ler expression overcomes repression of LEE in an sspA mutant. The expressions of virulence genes in wild type EHEC EDL933 (lane 1), the sspA mutant harboring the empty vector pACYC184 (pVector) (lane 2) and pACYC ler (p ler ) expressing ler from its native promoters (lane 3) were determined by primer extension analyses using labeled DNA oligos specific to LEE1/ler ( A ) , LEE2/espZ ( B ), LEE4/sepL ( C ), grlRA ( D ) and stcE ( E ) along with the ompA- specific oligo as a control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Figure Legend Snippet: Increased ler expression overcomes repression of LEE in an sspA mutant. The expressions of virulence genes in wild type EHEC EDL933 (lane 1), the sspA mutant harboring the empty vector pACYC184 (pVector) (lane 2) and pACYC ler (p ler ) expressing ler from its native promoters (lane 3) were determined by primer extension analyses using labeled DNA oligos specific to LEE1/ler ( A ) , LEE2/espZ ( B ), LEE4/sepL ( C ), grlRA ( D ) and stcE ( E ) along with the ompA- specific oligo as a control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Techniques Used: Expressing, Mutagenesis, Plasmid Preparation, Labeling

    SspA negatively affects H-NS levels in EHEC. The levels of H-NS were determined in wild type (lane 1) and sspA mutant (lane 2) derivatives of EHEC EDL933 grown to stationary phase cells by western blot. Equal amounts of total protein were resolved on a 10% Bis-Tris SDS-PAGE gel and transferred to a nitrocellulose membrane. Levels of H-NS and Fis were detected using polyclonal antibodies against the respective proteins. Fis served as an internal control for total protein levels. The relative amounts of H-NS normalized to that of Fis are indicated by the numbers in parenthesis.
    Figure Legend Snippet: SspA negatively affects H-NS levels in EHEC. The levels of H-NS were determined in wild type (lane 1) and sspA mutant (lane 2) derivatives of EHEC EDL933 grown to stationary phase cells by western blot. Equal amounts of total protein were resolved on a 10% Bis-Tris SDS-PAGE gel and transferred to a nitrocellulose membrane. Levels of H-NS and Fis were detected using polyclonal antibodies against the respective proteins. Fis served as an internal control for total protein levels. The relative amounts of H-NS normalized to that of Fis are indicated by the numbers in parenthesis.

    Techniques Used: Mutagenesis, Western Blot, SDS Page

    SspA is upstream of H-NS in the regulatory network of virulence gene expression in EHEC. The expression of virulence genes in wild type EHEC EDL933 (lane 1), sspA (lane 2) , hns (lane 3) and hns sspA (lane 4) mutant derivatives was determined by primer extension analyses using labeled DNA oligos specific to the transcripts of LEE1/ler ( A ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ). In each reaction, the ompA transcript served as an internal control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Figure Legend Snippet: SspA is upstream of H-NS in the regulatory network of virulence gene expression in EHEC. The expression of virulence genes in wild type EHEC EDL933 (lane 1), sspA (lane 2) , hns (lane 3) and hns sspA (lane 4) mutant derivatives was determined by primer extension analyses using labeled DNA oligos specific to the transcripts of LEE1/ler ( A ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ). In each reaction, the ompA transcript served as an internal control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Techniques Used: Expressing, Mutagenesis, Labeling

    SspA is required for cell adherence and A/E lesion formation. HEp-2 cells were infected by wild type EHEC EDL933 ( A ) and its mutant derivatives of sspA ( B ), sspA pQE sspA ( C ), sspA pQE sspA84-86 ( D ), hns ( E ) and hns sspA ( F ). Bacterial adherence was examined by phase-contrast images (left panels) and the actin cytoskeleton of infected HEp-2 cells by fluorescent microscopic images (right panels). Representative images are shown. Black and white arrowheads indicate bacteria and A/E lesions, respectively.
    Figure Legend Snippet: SspA is required for cell adherence and A/E lesion formation. HEp-2 cells were infected by wild type EHEC EDL933 ( A ) and its mutant derivatives of sspA ( B ), sspA pQE sspA ( C ), sspA pQE sspA84-86 ( D ), hns ( E ) and hns sspA ( F ). Bacterial adherence was examined by phase-contrast images (left panels) and the actin cytoskeleton of infected HEp-2 cells by fluorescent microscopic images (right panels). Representative images are shown. Black and white arrowheads indicate bacteria and A/E lesions, respectively.

    Techniques Used: Infection, Mutagenesis

    virus strains ehec o157 h7 edl933 atcc 43895 bl21 de3  (ATCC)


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    ATCC virus strains ehec o157 h7 edl933 atcc 43895 bl21 de3
    Virus Strains Ehec O157 H7 Edl933 Atcc 43895 Bl21 De3, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ehec o157 h7 edl933 strain atcc 700927  (ATCC)


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    ATCC ehec o157 h7 edl933 strain atcc 700927
    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC <t>EDL933</t> (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Ehec O157 H7 Edl933 Strain Atcc 700927, supplied by ATCC, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ehec o157 h7 edl933 strain atcc 700927/product/ATCC
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    ehec o157 h7 edl933 strain atcc 700927 - by Bioz Stars, 2024-02
    95/100 stars

    Images

    1) Product Images from "SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli"

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-12-231

    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC EDL933 (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Figure Legend Snippet: SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC EDL933 (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Techniques Used: Expressing, Mutagenesis, Labeling

    Increased ler expression overcomes repression of LEE in an sspA mutant. The expressions of virulence genes in wild type EHEC EDL933 (lane 1), the sspA mutant harboring the empty vector pACYC184 (pVector) (lane 2) and pACYC ler (p ler ) expressing ler from its native promoters (lane 3) were determined by primer extension analyses using labeled DNA oligos specific to LEE1/ler ( A ) , LEE2/espZ ( B ), LEE4/sepL ( C ), grlRA ( D ) and stcE ( E ) along with the ompA- specific oligo as a control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Figure Legend Snippet: Increased ler expression overcomes repression of LEE in an sspA mutant. The expressions of virulence genes in wild type EHEC EDL933 (lane 1), the sspA mutant harboring the empty vector pACYC184 (pVector) (lane 2) and pACYC ler (p ler ) expressing ler from its native promoters (lane 3) were determined by primer extension analyses using labeled DNA oligos specific to LEE1/ler ( A ) , LEE2/espZ ( B ), LEE4/sepL ( C ), grlRA ( D ) and stcE ( E ) along with the ompA- specific oligo as a control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Techniques Used: Expressing, Mutagenesis, Plasmid Preparation, Labeling

    SspA negatively affects H-NS levels in EHEC. The levels of H-NS were determined in wild type (lane 1) and sspA mutant (lane 2) derivatives of EHEC EDL933 grown to stationary phase cells by western blot. Equal amounts of total protein were resolved on a 10% Bis-Tris SDS-PAGE gel and transferred to a nitrocellulose membrane. Levels of H-NS and Fis were detected using polyclonal antibodies against the respective proteins. Fis served as an internal control for total protein levels. The relative amounts of H-NS normalized to that of Fis are indicated by the numbers in parenthesis.
    Figure Legend Snippet: SspA negatively affects H-NS levels in EHEC. The levels of H-NS were determined in wild type (lane 1) and sspA mutant (lane 2) derivatives of EHEC EDL933 grown to stationary phase cells by western blot. Equal amounts of total protein were resolved on a 10% Bis-Tris SDS-PAGE gel and transferred to a nitrocellulose membrane. Levels of H-NS and Fis were detected using polyclonal antibodies against the respective proteins. Fis served as an internal control for total protein levels. The relative amounts of H-NS normalized to that of Fis are indicated by the numbers in parenthesis.

    Techniques Used: Mutagenesis, Western Blot, SDS Page

    SspA is upstream of H-NS in the regulatory network of virulence gene expression in EHEC. The expression of virulence genes in wild type EHEC EDL933 (lane 1), sspA (lane 2) , hns (lane 3) and hns sspA (lane 4) mutant derivatives was determined by primer extension analyses using labeled DNA oligos specific to the transcripts of LEE1/ler ( A ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ). In each reaction, the ompA transcript served as an internal control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Figure Legend Snippet: SspA is upstream of H-NS in the regulatory network of virulence gene expression in EHEC. The expression of virulence genes in wild type EHEC EDL933 (lane 1), sspA (lane 2) , hns (lane 3) and hns sspA (lane 4) mutant derivatives was determined by primer extension analyses using labeled DNA oligos specific to the transcripts of LEE1/ler ( A ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ). In each reaction, the ompA transcript served as an internal control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Techniques Used: Expressing, Mutagenesis, Labeling

    SspA is required for cell adherence and A/E lesion formation. HEp-2 cells were infected by wild type EHEC EDL933 ( A ) and its mutant derivatives of sspA ( B ), sspA pQE sspA ( C ), sspA pQE sspA84-86 ( D ), hns ( E ) and hns sspA ( F ). Bacterial adherence was examined by phase-contrast images (left panels) and the actin cytoskeleton of infected HEp-2 cells by fluorescent microscopic images (right panels). Representative images are shown. Black and white arrowheads indicate bacteria and A/E lesions, respectively.
    Figure Legend Snippet: SspA is required for cell adherence and A/E lesion formation. HEp-2 cells were infected by wild type EHEC EDL933 ( A ) and its mutant derivatives of sspA ( B ), sspA pQE sspA ( C ), sspA pQE sspA84-86 ( D ), hns ( E ) and hns sspA ( F ). Bacterial adherence was examined by phase-contrast images (left panels) and the actin cytoskeleton of infected HEp-2 cells by fluorescent microscopic images (right panels). Representative images are shown. Black and white arrowheads indicate bacteria and A/E lesions, respectively.

    Techniques Used: Infection, Mutagenesis

    ehec edl933  (ATCC)


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    ATCC ehec edl933
    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC <t>EDL933</t> (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Ehec Edl933, supplied by ATCC, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ehec edl933/product/ATCC
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    ehec edl933 - by Bioz Stars, 2024-02
    95/100 stars

    Images

    1) Product Images from "SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli"

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-12-231

    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC EDL933 (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Figure Legend Snippet: SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC EDL933 (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Techniques Used: Expressing, Mutagenesis, Labeling

    Increased ler expression overcomes repression of LEE in an sspA mutant. The expressions of virulence genes in wild type EHEC EDL933 (lane 1), the sspA mutant harboring the empty vector pACYC184 (pVector) (lane 2) and pACYC ler (p ler ) expressing ler from its native promoters (lane 3) were determined by primer extension analyses using labeled DNA oligos specific to LEE1/ler ( A ) , LEE2/espZ ( B ), LEE4/sepL ( C ), grlRA ( D ) and stcE ( E ) along with the ompA- specific oligo as a control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Figure Legend Snippet: Increased ler expression overcomes repression of LEE in an sspA mutant. The expressions of virulence genes in wild type EHEC EDL933 (lane 1), the sspA mutant harboring the empty vector pACYC184 (pVector) (lane 2) and pACYC ler (p ler ) expressing ler from its native promoters (lane 3) were determined by primer extension analyses using labeled DNA oligos specific to LEE1/ler ( A ) , LEE2/espZ ( B ), LEE4/sepL ( C ), grlRA ( D ) and stcE ( E ) along with the ompA- specific oligo as a control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Techniques Used: Expressing, Mutagenesis, Plasmid Preparation, Labeling

    SspA negatively affects H-NS levels in EHEC. The levels of H-NS were determined in wild type (lane 1) and sspA mutant (lane 2) derivatives of EHEC EDL933 grown to stationary phase cells by western blot. Equal amounts of total protein were resolved on a 10% Bis-Tris SDS-PAGE gel and transferred to a nitrocellulose membrane. Levels of H-NS and Fis were detected using polyclonal antibodies against the respective proteins. Fis served as an internal control for total protein levels. The relative amounts of H-NS normalized to that of Fis are indicated by the numbers in parenthesis.
    Figure Legend Snippet: SspA negatively affects H-NS levels in EHEC. The levels of H-NS were determined in wild type (lane 1) and sspA mutant (lane 2) derivatives of EHEC EDL933 grown to stationary phase cells by western blot. Equal amounts of total protein were resolved on a 10% Bis-Tris SDS-PAGE gel and transferred to a nitrocellulose membrane. Levels of H-NS and Fis were detected using polyclonal antibodies against the respective proteins. Fis served as an internal control for total protein levels. The relative amounts of H-NS normalized to that of Fis are indicated by the numbers in parenthesis.

    Techniques Used: Mutagenesis, Western Blot, SDS Page

    SspA is upstream of H-NS in the regulatory network of virulence gene expression in EHEC. The expression of virulence genes in wild type EHEC EDL933 (lane 1), sspA (lane 2) , hns (lane 3) and hns sspA (lane 4) mutant derivatives was determined by primer extension analyses using labeled DNA oligos specific to the transcripts of LEE1/ler ( A ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ). In each reaction, the ompA transcript served as an internal control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Figure Legend Snippet: SspA is upstream of H-NS in the regulatory network of virulence gene expression in EHEC. The expression of virulence genes in wild type EHEC EDL933 (lane 1), sspA (lane 2) , hns (lane 3) and hns sspA (lane 4) mutant derivatives was determined by primer extension analyses using labeled DNA oligos specific to the transcripts of LEE1/ler ( A ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ). In each reaction, the ompA transcript served as an internal control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Techniques Used: Expressing, Mutagenesis, Labeling

    SspA is required for cell adherence and A/E lesion formation. HEp-2 cells were infected by wild type EHEC EDL933 ( A ) and its mutant derivatives of sspA ( B ), sspA pQE sspA ( C ), sspA pQE sspA84-86 ( D ), hns ( E ) and hns sspA ( F ). Bacterial adherence was examined by phase-contrast images (left panels) and the actin cytoskeleton of infected HEp-2 cells by fluorescent microscopic images (right panels). Representative images are shown. Black and white arrowheads indicate bacteria and A/E lesions, respectively.
    Figure Legend Snippet: SspA is required for cell adherence and A/E lesion formation. HEp-2 cells were infected by wild type EHEC EDL933 ( A ) and its mutant derivatives of sspA ( B ), sspA pQE sspA ( C ), sspA pQE sspA84-86 ( D ), hns ( E ) and hns sspA ( F ). Bacterial adherence was examined by phase-contrast images (left panels) and the actin cytoskeleton of infected HEp-2 cells by fluorescent microscopic images (right panels). Representative images are shown. Black and white arrowheads indicate bacteria and A/E lesions, respectively.

    Techniques Used: Infection, Mutagenesis

    enterohaemorrhagic e coli ehec strain edl933  (ATCC)


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    ATCC enterohaemorrhagic e coli ehec strain edl933
    Typical IS3 profiling gel . Lanes 1 and 29: 1 Kb ladder plus (Invitrogen); Lanes 2-12: EAEC strains AA 60A, NA H191-1, AA H232-1, AA 17-2, AA 253-1, AA 6-1, AA DS65-R2, AA501-1, AA H223-1, DA WC212-11 and AA DS67-R2; Lanes 13-25: AA H38-1, AA 042, AA 144-1, AA 44-1, AA H145-1, AA 309-1, AA 103-1, DA H92-1, AA 435-1, AA 199-1, AA H194-2, AA 278-1 and AA 239-1; Lanes 26-28: Control strains EHEC O157 <t>EDL933,</t> Shigella flexneri 2a 2425T and E. coli K-12 MG1655. Boxed numbers indicate bands described in Table 1.
    Enterohaemorrhagic E Coli Ehec Strain Edl933, supplied by ATCC, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "IS3 profiling identifies the enterohaemorrhagic Escherichia coli O-island 62 in a distinct enteroaggregative E. coli lineage"

    Article Title: IS3 profiling identifies the enterohaemorrhagic Escherichia coli O-island 62 in a distinct enteroaggregative E. coli lineage

    Journal: Gut Pathogens

    doi: 10.1186/1757-4749-3-4

    Typical IS3 profiling gel . Lanes 1 and 29: 1 Kb ladder plus (Invitrogen); Lanes 2-12: EAEC strains AA 60A, NA H191-1, AA H232-1, AA 17-2, AA 253-1, AA 6-1, AA DS65-R2, AA501-1, AA H223-1, DA WC212-11 and AA DS67-R2; Lanes 13-25: AA H38-1, AA 042, AA 144-1, AA 44-1, AA H145-1, AA 309-1, AA 103-1, DA H92-1, AA 435-1, AA 199-1, AA H194-2, AA 278-1 and AA 239-1; Lanes 26-28: Control strains EHEC O157 EDL933, Shigella flexneri 2a 2425T and E. coli K-12 MG1655. Boxed numbers indicate bands described in Table 1.
    Figure Legend Snippet: Typical IS3 profiling gel . Lanes 1 and 29: 1 Kb ladder plus (Invitrogen); Lanes 2-12: EAEC strains AA 60A, NA H191-1, AA H232-1, AA 17-2, AA 253-1, AA 6-1, AA DS65-R2, AA501-1, AA H223-1, DA WC212-11 and AA DS67-R2; Lanes 13-25: AA H38-1, AA 042, AA 144-1, AA 44-1, AA H145-1, AA 309-1, AA 103-1, DA H92-1, AA 435-1, AA 199-1, AA H194-2, AA 278-1 and AA 239-1; Lanes 26-28: Control strains EHEC O157 EDL933, Shigella flexneri 2a 2425T and E. coli K-12 MG1655. Boxed numbers indicate bands described in Table 1.

    Techniques Used:

    Genetic loci identified by IS3 profiling
    Figure Legend Snippet: Genetic loci identified by IS3 profiling

    Techniques Used: Selection, Sequencing, Diagnostic Assay, Plasmid Preparation

    EHEC O157:H7 strain EDL933 genome segment 2016573-2026572, containing O-island 62, and the corresponding regions in ECOR D EAEC strain 042 and E. coli K12 strain MG1655, illustrating the mosaic nature of the island .
    Figure Legend Snippet: EHEC O157:H7 strain EDL933 genome segment 2016573-2026572, containing O-island 62, and the corresponding regions in ECOR D EAEC strain 042 and E. coli K12 strain MG1655, illustrating the mosaic nature of the island .

    Techniques Used:

    ehec edl933  (ATCC)


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    ATCC ehec edl933
    Bacterial strains investigated in this study.
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    1) Product Images from "Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli"

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-7-21

    Bacterial strains investigated in this study.
    Figure Legend Snippet: Bacterial strains investigated in this study.

    Techniques Used:

    A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.
    Figure Legend Snippet: A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.

    Techniques Used: Mutagenesis, Incubation, Clone Assay, Plasmid Preparation, Positive Control, Transformation Assay

    Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.
    Figure Legend Snippet: Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.

    Techniques Used: Functional Assay, Mutagenesis

    Plasmids used in this study.
    Figure Legend Snippet: Plasmids used in this study.

    Techniques Used: Plasmid Preparation, Selection, Amplification, Clone Assay, Mutagenesis

    Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.
    Figure Legend Snippet: Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.

    Techniques Used: Construct, Plasmid Preparation, Amplification, Clone Assay, Mutagenesis, Transformation Assay, Electroporation

    ehec edl933  (ATCC)


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    ATCC ehec edl933
    Bacterial strains investigated in this study.
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    1) Product Images from "Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli"

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-7-21

    Bacterial strains investigated in this study.
    Figure Legend Snippet: Bacterial strains investigated in this study.

    Techniques Used:

    A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.
    Figure Legend Snippet: A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.

    Techniques Used: Mutagenesis, Incubation, Clone Assay, Plasmid Preparation, Positive Control, Transformation Assay

    Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.
    Figure Legend Snippet: Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.

    Techniques Used: Functional Assay, Mutagenesis

    Plasmids used in this study.
    Figure Legend Snippet: Plasmids used in this study.

    Techniques Used: Plasmid Preparation, Selection, Amplification, Clone Assay, Mutagenesis

    Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.
    Figure Legend Snippet: Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.

    Techniques Used: Construct, Plasmid Preparation, Amplification, Clone Assay, Mutagenesis, Transformation Assay, Electroporation

    ehec edl933  (ATCC)


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    ATCC ehec edl933
    Bacterial strains investigated in this study.
    Ehec Edl933, supplied by ATCC, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli"

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-7-21

    Bacterial strains investigated in this study.
    Figure Legend Snippet: Bacterial strains investigated in this study.

    Techniques Used:

    A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.
    Figure Legend Snippet: A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.

    Techniques Used: Mutagenesis, Incubation, Clone Assay, Plasmid Preparation, Positive Control, Transformation Assay

    Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.
    Figure Legend Snippet: Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.

    Techniques Used: Functional Assay, Mutagenesis

    Plasmids used in this study.
    Figure Legend Snippet: Plasmids used in this study.

    Techniques Used: Plasmid Preparation, Selection, Amplification, Clone Assay, Mutagenesis

    Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.
    Figure Legend Snippet: Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.

    Techniques Used: Construct, Plasmid Preparation, Amplification, Clone Assay, Mutagenesis, Transformation Assay, Electroporation

    ehec edl933 b  (ATCC)


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    ATCC ehec edl933 b
    Bacterial strains investigated in this study.
    Ehec Edl933 B, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli"

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-7-21

    Bacterial strains investigated in this study.
    Figure Legend Snippet: Bacterial strains investigated in this study.

    Techniques Used:

    A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.
    Figure Legend Snippet: A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.

    Techniques Used: Mutagenesis, Incubation, Clone Assay, Plasmid Preparation, Positive Control, Transformation Assay

    Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.
    Figure Legend Snippet: Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.

    Techniques Used: Functional Assay, Mutagenesis

    Plasmids used in this study.
    Figure Legend Snippet: Plasmids used in this study.

    Techniques Used: Plasmid Preparation, Selection, Amplification, Clone Assay, Mutagenesis

    Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.
    Figure Legend Snippet: Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.

    Techniques Used: Construct, Plasmid Preparation, Amplification, Clone Assay, Mutagenesis, Transformation Assay, Electroporation

    prototype o157 h7 ehec edl933  (ATCC)


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    ATCC prototype o157 h7 ehec edl933
    Bacterial strains investigated in this study.
    Prototype O157 H7 Ehec Edl933, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli"

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-7-21

    Bacterial strains investigated in this study.
    Figure Legend Snippet: Bacterial strains investigated in this study.

    Techniques Used:

    A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.
    Figure Legend Snippet: A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.

    Techniques Used: Mutagenesis, Incubation, Clone Assay, Plasmid Preparation, Positive Control, Transformation Assay

    Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.
    Figure Legend Snippet: Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.

    Techniques Used: Functional Assay, Mutagenesis

    Plasmids used in this study.
    Figure Legend Snippet: Plasmids used in this study.

    Techniques Used: Plasmid Preparation, Selection, Amplification, Clone Assay, Mutagenesis

    Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.
    Figure Legend Snippet: Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.

    Techniques Used: Construct, Plasmid Preparation, Amplification, Clone Assay, Mutagenesis, Transformation Assay, Electroporation

    ehec edl933  (ATCC)


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    Structured Review

    ATCC ehec edl933
    Bacterial strains investigated in this study.
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    1) Product Images from "Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli"

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-7-21

    Bacterial strains investigated in this study.
    Figure Legend Snippet: Bacterial strains investigated in this study.

    Techniques Used:

    A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.
    Figure Legend Snippet: A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.

    Techniques Used: Mutagenesis, Incubation, Clone Assay, Plasmid Preparation, Positive Control, Transformation Assay

    Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.
    Figure Legend Snippet: Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.

    Techniques Used: Functional Assay, Mutagenesis

    Plasmids used in this study.
    Figure Legend Snippet: Plasmids used in this study.

    Techniques Used: Plasmid Preparation, Selection, Amplification, Clone Assay, Mutagenesis

    Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.
    Figure Legend Snippet: Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.

    Techniques Used: Construct, Plasmid Preparation, Amplification, Clone Assay, Mutagenesis, Transformation Assay, Electroporation

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    ATCC ehec o157 h7 edl933 strain atcc 700927
    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC <t>EDL933</t> (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Ehec O157 H7 Edl933 Strain Atcc 700927, supplied by ATCC, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC virus strains ehec o157 h7 edl933 atcc 43895 bl21 de3
    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC <t>EDL933</t> (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
    Virus Strains Ehec O157 H7 Edl933 Atcc 43895 Bl21 De3, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC ehec edl933
    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC <t>EDL933</t> (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.
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    ATCC enterohaemorrhagic e coli ehec strain edl933
    Typical IS3 profiling gel . Lanes 1 and 29: 1 Kb ladder plus (Invitrogen); Lanes 2-12: EAEC strains AA 60A, NA H191-1, AA H232-1, AA 17-2, AA 253-1, AA 6-1, AA DS65-R2, AA501-1, AA H223-1, DA WC212-11 and AA DS67-R2; Lanes 13-25: AA H38-1, AA 042, AA 144-1, AA 44-1, AA H145-1, AA 309-1, AA 103-1, DA H92-1, AA 435-1, AA 199-1, AA H194-2, AA 278-1 and AA 239-1; Lanes 26-28: Control strains EHEC O157 <t>EDL933,</t> Shigella flexneri 2a 2425T and E. coli K-12 MG1655. Boxed numbers indicate bands described in Table 1.
    Enterohaemorrhagic E Coli Ehec Strain Edl933, supplied by ATCC, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC ehec edl933 b
    Bacterial strains investigated in this study.
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    ATCC prototype o157 h7 ehec edl933
    Bacterial strains investigated in this study.
    Prototype O157 H7 Ehec Edl933, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC EDL933 (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Journal: BMC Microbiology

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    doi: 10.1186/1471-2180-12-231

    Figure Lengend Snippet: SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC EDL933 (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Article Snippet: Gene deletions were constructed in EHEC O157:H7 EDL933 strain ATCC 700927 (Perna et al. 2001) by Lambda Red-mediated recombination using linear DNA fragments as described [ ].

    Techniques: Expressing, Mutagenesis, Labeling

    Increased ler expression overcomes repression of LEE in an sspA mutant. The expressions of virulence genes in wild type EHEC EDL933 (lane 1), the sspA mutant harboring the empty vector pACYC184 (pVector) (lane 2) and pACYC ler (p ler ) expressing ler from its native promoters (lane 3) were determined by primer extension analyses using labeled DNA oligos specific to LEE1/ler ( A ) , LEE2/espZ ( B ), LEE4/sepL ( C ), grlRA ( D ) and stcE ( E ) along with the ompA- specific oligo as a control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Journal: BMC Microbiology

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    doi: 10.1186/1471-2180-12-231

    Figure Lengend Snippet: Increased ler expression overcomes repression of LEE in an sspA mutant. The expressions of virulence genes in wild type EHEC EDL933 (lane 1), the sspA mutant harboring the empty vector pACYC184 (pVector) (lane 2) and pACYC ler (p ler ) expressing ler from its native promoters (lane 3) were determined by primer extension analyses using labeled DNA oligos specific to LEE1/ler ( A ) , LEE2/espZ ( B ), LEE4/sepL ( C ), grlRA ( D ) and stcE ( E ) along with the ompA- specific oligo as a control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Article Snippet: Gene deletions were constructed in EHEC O157:H7 EDL933 strain ATCC 700927 (Perna et al. 2001) by Lambda Red-mediated recombination using linear DNA fragments as described [ ].

    Techniques: Expressing, Mutagenesis, Plasmid Preparation, Labeling

    SspA negatively affects H-NS levels in EHEC. The levels of H-NS were determined in wild type (lane 1) and sspA mutant (lane 2) derivatives of EHEC EDL933 grown to stationary phase cells by western blot. Equal amounts of total protein were resolved on a 10% Bis-Tris SDS-PAGE gel and transferred to a nitrocellulose membrane. Levels of H-NS and Fis were detected using polyclonal antibodies against the respective proteins. Fis served as an internal control for total protein levels. The relative amounts of H-NS normalized to that of Fis are indicated by the numbers in parenthesis.

    Journal: BMC Microbiology

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    doi: 10.1186/1471-2180-12-231

    Figure Lengend Snippet: SspA negatively affects H-NS levels in EHEC. The levels of H-NS were determined in wild type (lane 1) and sspA mutant (lane 2) derivatives of EHEC EDL933 grown to stationary phase cells by western blot. Equal amounts of total protein were resolved on a 10% Bis-Tris SDS-PAGE gel and transferred to a nitrocellulose membrane. Levels of H-NS and Fis were detected using polyclonal antibodies against the respective proteins. Fis served as an internal control for total protein levels. The relative amounts of H-NS normalized to that of Fis are indicated by the numbers in parenthesis.

    Article Snippet: Gene deletions were constructed in EHEC O157:H7 EDL933 strain ATCC 700927 (Perna et al. 2001) by Lambda Red-mediated recombination using linear DNA fragments as described [ ].

    Techniques: Mutagenesis, Western Blot, SDS Page

    SspA is upstream of H-NS in the regulatory network of virulence gene expression in EHEC. The expression of virulence genes in wild type EHEC EDL933 (lane 1), sspA (lane 2) , hns (lane 3) and hns sspA (lane 4) mutant derivatives was determined by primer extension analyses using labeled DNA oligos specific to the transcripts of LEE1/ler ( A ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ). In each reaction, the ompA transcript served as an internal control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Journal: BMC Microbiology

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    doi: 10.1186/1471-2180-12-231

    Figure Lengend Snippet: SspA is upstream of H-NS in the regulatory network of virulence gene expression in EHEC. The expression of virulence genes in wild type EHEC EDL933 (lane 1), sspA (lane 2) , hns (lane 3) and hns sspA (lane 4) mutant derivatives was determined by primer extension analyses using labeled DNA oligos specific to the transcripts of LEE1/ler ( A ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ). In each reaction, the ompA transcript served as an internal control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Article Snippet: Gene deletions were constructed in EHEC O157:H7 EDL933 strain ATCC 700927 (Perna et al. 2001) by Lambda Red-mediated recombination using linear DNA fragments as described [ ].

    Techniques: Expressing, Mutagenesis, Labeling

    SspA is required for cell adherence and A/E lesion formation. HEp-2 cells were infected by wild type EHEC EDL933 ( A ) and its mutant derivatives of sspA ( B ), sspA pQE sspA ( C ), sspA pQE sspA84-86 ( D ), hns ( E ) and hns sspA ( F ). Bacterial adherence was examined by phase-contrast images (left panels) and the actin cytoskeleton of infected HEp-2 cells by fluorescent microscopic images (right panels). Representative images are shown. Black and white arrowheads indicate bacteria and A/E lesions, respectively.

    Journal: BMC Microbiology

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    doi: 10.1186/1471-2180-12-231

    Figure Lengend Snippet: SspA is required for cell adherence and A/E lesion formation. HEp-2 cells were infected by wild type EHEC EDL933 ( A ) and its mutant derivatives of sspA ( B ), sspA pQE sspA ( C ), sspA pQE sspA84-86 ( D ), hns ( E ) and hns sspA ( F ). Bacterial adherence was examined by phase-contrast images (left panels) and the actin cytoskeleton of infected HEp-2 cells by fluorescent microscopic images (right panels). Representative images are shown. Black and white arrowheads indicate bacteria and A/E lesions, respectively.

    Article Snippet: Gene deletions were constructed in EHEC O157:H7 EDL933 strain ATCC 700927 (Perna et al. 2001) by Lambda Red-mediated recombination using linear DNA fragments as described [ ].

    Techniques: Infection, Mutagenesis

    SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC EDL933 (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Journal: BMC Microbiology

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    doi: 10.1186/1471-2180-12-231

    Figure Lengend Snippet: SspA positively affects LEE expression in stationary phase cells. Primer extension analyses on total RNA extracted from wild type EHEC EDL933 (lane 1), the sspA mutant (lane 2) and the sspA mutant complemented with wild type sspA (lane 3) or mutant sspA84-86 (lane 4) as indicated, grown in LB at 37°C to stationary phase (OD 600 ~ 3.0). The Labeled DNA oligos specific to the transcripts of LEE1/ler ( A and I ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ) were used. The ompA transcripts, detected with a labeled ompA -specific DNA oligo, served as internal control for the primer extension reaction. Wild type and mutant SspA were expressed from pQE sspA and pQE sspA84-86 respectively in the absence of induction at similar levels. The transcripts LEE1-5, map, grlRA, stcE and the control transcript ompA are indicated. The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Article Snippet: The ability of EHEC EDL933 (ATCC 700927) wild type and its mutant derivatives to adhere and form A/E lesions on HEp-2 cell monolayers was evaluated using the fluorescent actin staining assay as described [ ].

    Techniques: Expressing, Mutagenesis, Labeling

    Increased ler expression overcomes repression of LEE in an sspA mutant. The expressions of virulence genes in wild type EHEC EDL933 (lane 1), the sspA mutant harboring the empty vector pACYC184 (pVector) (lane 2) and pACYC ler (p ler ) expressing ler from its native promoters (lane 3) were determined by primer extension analyses using labeled DNA oligos specific to LEE1/ler ( A ) , LEE2/espZ ( B ), LEE4/sepL ( C ), grlRA ( D ) and stcE ( E ) along with the ompA- specific oligo as a control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Journal: BMC Microbiology

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    doi: 10.1186/1471-2180-12-231

    Figure Lengend Snippet: Increased ler expression overcomes repression of LEE in an sspA mutant. The expressions of virulence genes in wild type EHEC EDL933 (lane 1), the sspA mutant harboring the empty vector pACYC184 (pVector) (lane 2) and pACYC ler (p ler ) expressing ler from its native promoters (lane 3) were determined by primer extension analyses using labeled DNA oligos specific to LEE1/ler ( A ) , LEE2/espZ ( B ), LEE4/sepL ( C ), grlRA ( D ) and stcE ( E ) along with the ompA- specific oligo as a control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Article Snippet: The ability of EHEC EDL933 (ATCC 700927) wild type and its mutant derivatives to adhere and form A/E lesions on HEp-2 cell monolayers was evaluated using the fluorescent actin staining assay as described [ ].

    Techniques: Expressing, Mutagenesis, Plasmid Preparation, Labeling

    SspA negatively affects H-NS levels in EHEC. The levels of H-NS were determined in wild type (lane 1) and sspA mutant (lane 2) derivatives of EHEC EDL933 grown to stationary phase cells by western blot. Equal amounts of total protein were resolved on a 10% Bis-Tris SDS-PAGE gel and transferred to a nitrocellulose membrane. Levels of H-NS and Fis were detected using polyclonal antibodies against the respective proteins. Fis served as an internal control for total protein levels. The relative amounts of H-NS normalized to that of Fis are indicated by the numbers in parenthesis.

    Journal: BMC Microbiology

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    doi: 10.1186/1471-2180-12-231

    Figure Lengend Snippet: SspA negatively affects H-NS levels in EHEC. The levels of H-NS were determined in wild type (lane 1) and sspA mutant (lane 2) derivatives of EHEC EDL933 grown to stationary phase cells by western blot. Equal amounts of total protein were resolved on a 10% Bis-Tris SDS-PAGE gel and transferred to a nitrocellulose membrane. Levels of H-NS and Fis were detected using polyclonal antibodies against the respective proteins. Fis served as an internal control for total protein levels. The relative amounts of H-NS normalized to that of Fis are indicated by the numbers in parenthesis.

    Article Snippet: The ability of EHEC EDL933 (ATCC 700927) wild type and its mutant derivatives to adhere and form A/E lesions on HEp-2 cell monolayers was evaluated using the fluorescent actin staining assay as described [ ].

    Techniques: Mutagenesis, Western Blot, SDS Page

    SspA is upstream of H-NS in the regulatory network of virulence gene expression in EHEC. The expression of virulence genes in wild type EHEC EDL933 (lane 1), sspA (lane 2) , hns (lane 3) and hns sspA (lane 4) mutant derivatives was determined by primer extension analyses using labeled DNA oligos specific to the transcripts of LEE1/ler ( A ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ). In each reaction, the ompA transcript served as an internal control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Journal: BMC Microbiology

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    doi: 10.1186/1471-2180-12-231

    Figure Lengend Snippet: SspA is upstream of H-NS in the regulatory network of virulence gene expression in EHEC. The expression of virulence genes in wild type EHEC EDL933 (lane 1), sspA (lane 2) , hns (lane 3) and hns sspA (lane 4) mutant derivatives was determined by primer extension analyses using labeled DNA oligos specific to the transcripts of LEE1/ler ( A ) , LEE2/espZ ( B ) , LEE3/mpc ( C ) , LEE4/sepL ( D ), LEE5/tir ( E ), map ( F ), grlRA ( G ) and stcE ( H ). In each reaction, the ompA transcript served as an internal control. Samples were prepared and analyzed as described in the legend of Figure . The relative transcript levels of target genes normalized to that of ompA are indicated by the numbers in parenthesis.

    Article Snippet: The ability of EHEC EDL933 (ATCC 700927) wild type and its mutant derivatives to adhere and form A/E lesions on HEp-2 cell monolayers was evaluated using the fluorescent actin staining assay as described [ ].

    Techniques: Expressing, Mutagenesis, Labeling

    SspA is required for cell adherence and A/E lesion formation. HEp-2 cells were infected by wild type EHEC EDL933 ( A ) and its mutant derivatives of sspA ( B ), sspA pQE sspA ( C ), sspA pQE sspA84-86 ( D ), hns ( E ) and hns sspA ( F ). Bacterial adherence was examined by phase-contrast images (left panels) and the actin cytoskeleton of infected HEp-2 cells by fluorescent microscopic images (right panels). Representative images are shown. Black and white arrowheads indicate bacteria and A/E lesions, respectively.

    Journal: BMC Microbiology

    Article Title: SspA up-regulates gene expression of the LEE pathogenicity island by decreasing H-NS levels in enterohemorrhagic Escherichia coli

    doi: 10.1186/1471-2180-12-231

    Figure Lengend Snippet: SspA is required for cell adherence and A/E lesion formation. HEp-2 cells were infected by wild type EHEC EDL933 ( A ) and its mutant derivatives of sspA ( B ), sspA pQE sspA ( C ), sspA pQE sspA84-86 ( D ), hns ( E ) and hns sspA ( F ). Bacterial adherence was examined by phase-contrast images (left panels) and the actin cytoskeleton of infected HEp-2 cells by fluorescent microscopic images (right panels). Representative images are shown. Black and white arrowheads indicate bacteria and A/E lesions, respectively.

    Article Snippet: The ability of EHEC EDL933 (ATCC 700927) wild type and its mutant derivatives to adhere and form A/E lesions on HEp-2 cell monolayers was evaluated using the fluorescent actin staining assay as described [ ].

    Techniques: Infection, Mutagenesis

    Typical IS3 profiling gel . Lanes 1 and 29: 1 Kb ladder plus (Invitrogen); Lanes 2-12: EAEC strains AA 60A, NA H191-1, AA H232-1, AA 17-2, AA 253-1, AA 6-1, AA DS65-R2, AA501-1, AA H223-1, DA WC212-11 and AA DS67-R2; Lanes 13-25: AA H38-1, AA 042, AA 144-1, AA 44-1, AA H145-1, AA 309-1, AA 103-1, DA H92-1, AA 435-1, AA 199-1, AA H194-2, AA 278-1 and AA 239-1; Lanes 26-28: Control strains EHEC O157 EDL933, Shigella flexneri 2a 2425T and E. coli K-12 MG1655. Boxed numbers indicate bands described in Table 1.

    Journal: Gut Pathogens

    Article Title: IS3 profiling identifies the enterohaemorrhagic Escherichia coli O-island 62 in a distinct enteroaggregative E. coli lineage

    doi: 10.1186/1757-4749-3-4

    Figure Lengend Snippet: Typical IS3 profiling gel . Lanes 1 and 29: 1 Kb ladder plus (Invitrogen); Lanes 2-12: EAEC strains AA 60A, NA H191-1, AA H232-1, AA 17-2, AA 253-1, AA 6-1, AA DS65-R2, AA501-1, AA H223-1, DA WC212-11 and AA DS67-R2; Lanes 13-25: AA H38-1, AA 042, AA 144-1, AA 44-1, AA H145-1, AA 309-1, AA 103-1, DA H92-1, AA 435-1, AA 199-1, AA H194-2, AA 278-1 and AA 239-1; Lanes 26-28: Control strains EHEC O157 EDL933, Shigella flexneri 2a 2425T and E. coli K-12 MG1655. Boxed numbers indicate bands described in Table 1.

    Article Snippet: Non-EAEC E. coli strains that were used as negative controls were E. coli K-12 strain MG1655, enterohaemorrhagic E. coli (EHEC) strain EDL933 (ATCC 43895) [ ], diffusely adherent E. coli strains DA WC212-11 and DA H92-1, enteropathogenic E. coli strains E2348/69 and B171-8 [ , ], uropathogenic E. coli strain 536 [ ], as well as Shigella flexneri 2a strain 2457T [ ].

    Techniques:

    Genetic loci identified by IS3 profiling

    Journal: Gut Pathogens

    Article Title: IS3 profiling identifies the enterohaemorrhagic Escherichia coli O-island 62 in a distinct enteroaggregative E. coli lineage

    doi: 10.1186/1757-4749-3-4

    Figure Lengend Snippet: Genetic loci identified by IS3 profiling

    Article Snippet: Non-EAEC E. coli strains that were used as negative controls were E. coli K-12 strain MG1655, enterohaemorrhagic E. coli (EHEC) strain EDL933 (ATCC 43895) [ ], diffusely adherent E. coli strains DA WC212-11 and DA H92-1, enteropathogenic E. coli strains E2348/69 and B171-8 [ , ], uropathogenic E. coli strain 536 [ ], as well as Shigella flexneri 2a strain 2457T [ ].

    Techniques: Selection, Sequencing, Diagnostic Assay, Plasmid Preparation

    EHEC O157:H7 strain EDL933 genome segment 2016573-2026572, containing O-island 62, and the corresponding regions in ECOR D EAEC strain 042 and E. coli K12 strain MG1655, illustrating the mosaic nature of the island .

    Journal: Gut Pathogens

    Article Title: IS3 profiling identifies the enterohaemorrhagic Escherichia coli O-island 62 in a distinct enteroaggregative E. coli lineage

    doi: 10.1186/1757-4749-3-4

    Figure Lengend Snippet: EHEC O157:H7 strain EDL933 genome segment 2016573-2026572, containing O-island 62, and the corresponding regions in ECOR D EAEC strain 042 and E. coli K12 strain MG1655, illustrating the mosaic nature of the island .

    Article Snippet: Non-EAEC E. coli strains that were used as negative controls were E. coli K-12 strain MG1655, enterohaemorrhagic E. coli (EHEC) strain EDL933 (ATCC 43895) [ ], diffusely adherent E. coli strains DA WC212-11 and DA H92-1, enteropathogenic E. coli strains E2348/69 and B171-8 [ , ], uropathogenic E. coli strain 536 [ ], as well as Shigella flexneri 2a strain 2457T [ ].

    Techniques:

    Bacterial strains investigated in this study.

    Journal: BMC Microbiology

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    doi: 10.1186/1471-2180-7-21

    Figure Lengend Snippet: Bacterial strains investigated in this study.

    Article Snippet: Phenotypical characterization of EHEC EDL933 b , a further clone of EDL933, and complementation experiments clearly showed that the stop codon in rpoS* causally determined the stress phenotype of ATCC strain 700927.

    Techniques:

    A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.

    Journal: BMC Microbiology

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    doi: 10.1186/1471-2180-7-21

    Figure Lengend Snippet: A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.

    Article Snippet: Phenotypical characterization of EHEC EDL933 b , a further clone of EDL933, and complementation experiments clearly showed that the stop codon in rpoS* causally determined the stress phenotype of ATCC strain 700927.

    Techniques: Mutagenesis, Incubation, Clone Assay, Plasmid Preparation, Positive Control, Transformation Assay

    Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.

    Journal: BMC Microbiology

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    doi: 10.1186/1471-2180-7-21

    Figure Lengend Snippet: Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.

    Article Snippet: Phenotypical characterization of EHEC EDL933 b , a further clone of EDL933, and complementation experiments clearly showed that the stop codon in rpoS* causally determined the stress phenotype of ATCC strain 700927.

    Techniques: Functional Assay, Mutagenesis

    Plasmids used in this study.

    Journal: BMC Microbiology

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    doi: 10.1186/1471-2180-7-21

    Figure Lengend Snippet: Plasmids used in this study.

    Article Snippet: Phenotypical characterization of EHEC EDL933 b , a further clone of EDL933, and complementation experiments clearly showed that the stop codon in rpoS* causally determined the stress phenotype of ATCC strain 700927.

    Techniques: Plasmid Preparation, Selection, Amplification, Clone Assay, Mutagenesis

    Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.

    Journal: BMC Microbiology

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    doi: 10.1186/1471-2180-7-21

    Figure Lengend Snippet: Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.

    Article Snippet: Phenotypical characterization of EHEC EDL933 b , a further clone of EDL933, and complementation experiments clearly showed that the stop codon in rpoS* causally determined the stress phenotype of ATCC strain 700927.

    Techniques: Construct, Plasmid Preparation, Amplification, Clone Assay, Mutagenesis, Transformation Assay, Electroporation

    Bacterial strains investigated in this study.

    Journal: BMC Microbiology

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    doi: 10.1186/1471-2180-7-21

    Figure Lengend Snippet: Bacterial strains investigated in this study.

    Article Snippet: Our study was initially focused on analyzing the role of σ S in acid stress behavior of two different EHEC isolates used as model organisms in our laboratory: the sequenced prototype O157:H7 EHEC EDL933 (ATCC 700927) [ ] and a very well characterized EHEC O26:H11 isolate from a HUS patient [ , ].

    Techniques:

    A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.

    Journal: BMC Microbiology

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    doi: 10.1186/1471-2180-7-21

    Figure Lengend Snippet: A: Acid resistance of EHEC 126814 and E. coli MHH126-5 . Inducible acid resistance of EHEC wild type strain 126814 and its isogenic rpoS deletion mutant E. coli MHH126-5 was investigated after 2 h incubation in LB media at pH 2.5 or 1.5. The wild type strain showed a high level of acid resistance, which was induced from OD 600 0.7 of the preparatory culture. It reached up to 115 % survival at OD 600 2.5 of the starter culture, indicating that EHEC 126814 was able to grow under these conditions. In LB media with pH 1.5 up to 75 % of the inoculum survived. E. coli MHH126-5 was completely sensitive to acid treatment regardless of pH and OD 600 of the preparatory culture. Percentage survival figures in relation to OD 600 of one typical experiment are depicted. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point. B: Acid resistance of EHEC EDL933 a , EDL933 b and E. coli MHH933-5 . EHEC EDL933 b was very acid resistant in all experiments. However, between EHEC wild type strain EDL933 a and its mutant E. coli MHH933-5 no differences could be observed. Both isolates showed weak resistance under acidic growth conditions and showed a similar behavior in all other experiments. In contrast to EHEC 126814, acid resistance of the O157 isolates was induced at OD 600 1.2 of the starter culture. One typical experiment has been shown as a representation of all independent tests carried out. The means and standard deviations were calculated from three independent dilution series prepared at each individual measuring point. C: Acid resistance of E. coli MHH126-5 complemented with pSC1 . This figure depicts the inducible acid resistance in relation to OD 600 of one typical experiment. Acid resistance was assayed at pH 2.5 and 1.5. By complementation of rpoS deletion mutant E. coli MHH126-5 with pSC1, containing its own rpoS gene cloned into plasmid pBR322, a phenotype could be restored that was even more resistant to acid stress than wild type strain EHEC 126814. OD 600 of acid resistance induction was identical to values obtained with the wild type strain, shown in figure 1A. In order to compare growth conditions in LB media containing ampicillin, positive control EHEC 126814 had been transformed with plasmid pBR322. Compared to figure 1A, the antibiotic and/or pBR322 negatively influenced acid resistance of this strain. The percentage survival at pH 2.5 was below 90 %. One typical experiment is shown representative of independent tests. The means and standard deviations were calculated from three independent dilution series made at each individual measuring point.

    Article Snippet: Our study was initially focused on analyzing the role of σ S in acid stress behavior of two different EHEC isolates used as model organisms in our laboratory: the sequenced prototype O157:H7 EHEC EDL933 (ATCC 700927) [ ] and a very well characterized EHEC O26:H11 isolate from a HUS patient [ , ].

    Techniques: Mutagenesis, Incubation, Clone Assay, Plasmid Preparation, Positive Control, Transformation Assay

    Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.

    Journal: BMC Microbiology

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    doi: 10.1186/1471-2180-7-21

    Figure Lengend Snippet: Functional analysis of further rpoS genes by complementation of E. coli MHH126-5 . This figure shows resistance data of the rpoS deletion mutant E. coli MHH126-5 complemented with each of the plasmids pUD2 (pBR322 + rpoS EDL933b ), pUD4 (pBR322 + rpoS 86-24 ), pUD10 (pBR322 + rpoS 288597 ), pUD8 (pBR322 + rpoS E-D53 ), pUD6 (pBR322 + rpoS E-D68 ), pUD9 (pBR322 + rpoS E-D68 ) or pDS4 (pBR322 + rpoS EcN ) in comparison to the corresponding wild type EHEC strains EDL933 b , 86-24 and 288597, STEC isolates E-D53 and E-D68 as well as EcN. All wild type strains are indicated by black bars, the complemented mutants by grey ones. Plasmids pUD2 and pUD4, pUD10 and pUD8 conferred an acid resistance phenotype to the mutant, which was comparable to the corresponding parental strain. Interestingly, upon complementation with pUD6 and pUD9, bearing the rpoS gene of STEC E-D68, E. coli MHH126-5 became strongly pH resistant. This was in sharp contrast to the behavior of the STEC E-D68 wild type strain. A similar phenomenon was observed when the acid resistance test strain E. coli MHH126-5 was complemented with pDS4 containing rpoS of EcN. The means and standard deviations were calculated from three independent dilution series in this exemplary experiment.

    Article Snippet: Our study was initially focused on analyzing the role of σ S in acid stress behavior of two different EHEC isolates used as model organisms in our laboratory: the sequenced prototype O157:H7 EHEC EDL933 (ATCC 700927) [ ] and a very well characterized EHEC O26:H11 isolate from a HUS patient [ , ].

    Techniques: Functional Assay, Mutagenesis

    Plasmids used in this study.

    Journal: BMC Microbiology

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    doi: 10.1186/1471-2180-7-21

    Figure Lengend Snippet: Plasmids used in this study.

    Article Snippet: Our study was initially focused on analyzing the role of σ S in acid stress behavior of two different EHEC isolates used as model organisms in our laboratory: the sequenced prototype O157:H7 EHEC EDL933 (ATCC 700927) [ ] and a very well characterized EHEC O26:H11 isolate from a HUS patient [ , ].

    Techniques: Plasmid Preparation, Selection, Amplification, Clone Assay, Mutagenesis

    Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.

    Journal: BMC Microbiology

    Article Title: Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

    doi: 10.1186/1471-2180-7-21

    Figure Lengend Snippet: Construction of complementation plasmids . Complementation plasmids pSC1, pSC2, pUD2, pUD4, pUD8, pUD6, pUD9, pUD10 and pDS4 were constructed using low copy vector pBR322 as backbone. Each of them harbors one complete rpoS gene and the rpoS p promoter [39] from EHEC/STEC strains 126814 (pSC1), EDL933 a (pSC2), EDL933 b (pUD2), 86-24 (pUD4), E-D53 (pUD8) and 288597 (pUD10), amplified by PCR with primers RpoS 8/RpoS 9 containing 5' Hind III restriction sites. RpoS 8 and RpoS 4c were taken to synthesize the respective DNA fragment from STEC E-D68 in order to generate the two independent plasmids pUD6 and pUD9. pDS4 contained the cloned rpoS gene from EcN, amplified with primers RpoS 8 and RpoS 22. pMH33 was made from pSC2 by PCR mediated site specific mutagenesis, employing the mutation rpoS T721G to resolve the TAA stop codon in rpoS *. RpoS negative mutant strain E. coli MHH126-5 was transformed with each of the complementation vectors by electroporation.

    Article Snippet: Our study was initially focused on analyzing the role of σ S in acid stress behavior of two different EHEC isolates used as model organisms in our laboratory: the sequenced prototype O157:H7 EHEC EDL933 (ATCC 700927) [ ] and a very well characterized EHEC O26:H11 isolate from a HUS patient [ , ].

    Techniques: Construct, Plasmid Preparation, Amplification, Clone Assay, Mutagenesis, Transformation Assay, Electroporation