sspi hf  (New England Biolabs)


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    Name:
    SspI HF
    Description:
    SspI HF 5 000 units
    Catalog Number:
    r3132l
    Price:
    290
    Category:
    Restriction Enzymes
    Size:
    5 000 units
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    Structured Review

    New England Biolabs sspi hf
    SspI HF
    SspI HF 5 000 units
    https://www.bioz.com/result/sspi hf/product/New England Biolabs
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sspi hf - by Bioz Stars, 2021-03
    93/100 stars

    Images

    1) Product Images from "Fallacy of the Unique Genome: Sequence Diversity within Single Helicobacter pylori Strains"

    Article Title: Fallacy of the Unique Genome: Sequence Diversity within Single Helicobacter pylori Strains

    Journal: mBio

    doi: 10.1128/mBio.02321-16

    H. pylori SS1 and PMSS1 are gene copy number variable at the cagA locus. (A) Schematic diagram showing tandem arrays of the identical 5,072-bp repeat regions at the cagA locus, all of which contain identical copies of the 3,540-bp cagA gene. The SspI fragment sizes of strains with 1 to 4 cagA copies are shown at the right. The drawing is not to scale. (B) Southern blot of SspI-digested genomic DNA from H. pylori SS1 or PMSS1 probed with a 297-bp PCR product amplified from SS1 cagA bp 1217 to 1514. The original working stocks of SS1 (lane 1) and PMSS1 (lane 5) showed bands corresponding in size to 4, 3, 2, and 1 copies of cagA (asterisks). Subculture of 4 single colonies from the freezer stocks demonstrated clones with 4, 3, or 2 copies of cagA (lanes 2 to 4) from SS1; subculture of 12 single colonies showed either 4 or 2 copies of cagA in PMSS1 (lanes 6 to 8). PMSS1 with a cagA deletion served as a negative control (lane 9). A kilobase ladder is shown at the left. (C) Western blot of H. pylori PMSS1 to examine whether the cagA copy number is positively correlated with protein expression. For this analysis, six individual single-colony isolates of PMSS1 were used, two each with four copies, two copies, or one copy of cagA . Relative quantities of protein in each band were determined using Image Lab software (Bio-Rad Laboratories). The density of the CagA band was divided by the density of the corresponding UreB band to obtain the normalized quantity of CagA to account for differences in gel loading.
    Figure Legend Snippet: H. pylori SS1 and PMSS1 are gene copy number variable at the cagA locus. (A) Schematic diagram showing tandem arrays of the identical 5,072-bp repeat regions at the cagA locus, all of which contain identical copies of the 3,540-bp cagA gene. The SspI fragment sizes of strains with 1 to 4 cagA copies are shown at the right. The drawing is not to scale. (B) Southern blot of SspI-digested genomic DNA from H. pylori SS1 or PMSS1 probed with a 297-bp PCR product amplified from SS1 cagA bp 1217 to 1514. The original working stocks of SS1 (lane 1) and PMSS1 (lane 5) showed bands corresponding in size to 4, 3, 2, and 1 copies of cagA (asterisks). Subculture of 4 single colonies from the freezer stocks demonstrated clones with 4, 3, or 2 copies of cagA (lanes 2 to 4) from SS1; subculture of 12 single colonies showed either 4 or 2 copies of cagA in PMSS1 (lanes 6 to 8). PMSS1 with a cagA deletion served as a negative control (lane 9). A kilobase ladder is shown at the left. (C) Western blot of H. pylori PMSS1 to examine whether the cagA copy number is positively correlated with protein expression. For this analysis, six individual single-colony isolates of PMSS1 were used, two each with four copies, two copies, or one copy of cagA . Relative quantities of protein in each band were determined using Image Lab software (Bio-Rad Laboratories). The density of the CagA band was divided by the density of the corresponding UreB band to obtain the normalized quantity of CagA to account for differences in gel loading.

    Techniques Used: Southern Blot, Polymerase Chain Reaction, Amplification, Clone Assay, Negative Control, Western Blot, Expressing, Software

    2) Product Images from "The HIV Protease Inhibitor Nelfinavir Inhibits Kaposi's Sarcoma-Associated Herpesvirus Replication In Vitro ▿"

    Article Title: The HIV Protease Inhibitor Nelfinavir Inhibits Kaposi's Sarcoma-Associated Herpesvirus Replication In Vitro ▿

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.01295-10

    Construction and quantitation of rKSHV.294. (A) Schematic diagram of rKSHV.294 showing the insertion site in the KSHV genome; the BamHI sites flanking the 4.8-kb segment of the KSHV genome used are indicated. The relative positions of the SeAP, the GFP, and the Neo elements are shown with their respective promoters but not to scale. Beneath the 294 construct are shown the expected PCR products (a to e) with the primers used for analysis of viral DNA. (B) Hybridization analysis of rKSHV.294 and JSC-1 viral DNA. (Left) KSHV probe using the 4.8-kb BamHI fragment used to construct the virus. (Right) Neo probe. Lane 1, rKSHV.294 × AflII, predicted fragments of 15.9, 5.7, and 4.8 kb. The 4.8-kb fragment contains the SeAP/GFP/Neo insert with only 170 bp of KSHV DNA, which accounts for the weak band. Lane 2, JSC-1 × AflII, predicted fragments of 15.9 and 5.8 kb. Lane 3, rKSHV.294 × SspI, predicted fragments of 8 and 6.2 kb. Lane 4, JSC-1 × SspI, predicted fragments of 6.2 and 3.3 kb. Lane 5, rKSHV.294 × AflII, predicted fragment of 4.8 kb. Lane 6, JSC-1 × AflII. Lane 7, rKSHV.294 DNA × SspI, predicted fragment of 8 kb. Lane 8, JSC-1 × SspI. DNA markers in kb are on the left side. (C) Ethidium bromide-stained agarose gel following electrophoresis of the PCR products resulting with rKSHV.294 viral DNA with the indicated primers described in Materials and Methods. Lane a, ORF56f (outside BamHI site) and SeAPr; lane b, ORF57u and SeAPr; lane c, SeAPf and GFPf; lane d, GFPr and Neor; lane e, Neof and K9. (D) rKSHV.294 was tested for tTA control of SeAP expression by the infection of 293 cells or of 293 cells expressing tTA at an MOI of
    Figure Legend Snippet: Construction and quantitation of rKSHV.294. (A) Schematic diagram of rKSHV.294 showing the insertion site in the KSHV genome; the BamHI sites flanking the 4.8-kb segment of the KSHV genome used are indicated. The relative positions of the SeAP, the GFP, and the Neo elements are shown with their respective promoters but not to scale. Beneath the 294 construct are shown the expected PCR products (a to e) with the primers used for analysis of viral DNA. (B) Hybridization analysis of rKSHV.294 and JSC-1 viral DNA. (Left) KSHV probe using the 4.8-kb BamHI fragment used to construct the virus. (Right) Neo probe. Lane 1, rKSHV.294 × AflII, predicted fragments of 15.9, 5.7, and 4.8 kb. The 4.8-kb fragment contains the SeAP/GFP/Neo insert with only 170 bp of KSHV DNA, which accounts for the weak band. Lane 2, JSC-1 × AflII, predicted fragments of 15.9 and 5.8 kb. Lane 3, rKSHV.294 × SspI, predicted fragments of 8 and 6.2 kb. Lane 4, JSC-1 × SspI, predicted fragments of 6.2 and 3.3 kb. Lane 5, rKSHV.294 × AflII, predicted fragment of 4.8 kb. Lane 6, JSC-1 × AflII. Lane 7, rKSHV.294 DNA × SspI, predicted fragment of 8 kb. Lane 8, JSC-1 × SspI. DNA markers in kb are on the left side. (C) Ethidium bromide-stained agarose gel following electrophoresis of the PCR products resulting with rKSHV.294 viral DNA with the indicated primers described in Materials and Methods. Lane a, ORF56f (outside BamHI site) and SeAPr; lane b, ORF57u and SeAPr; lane c, SeAPf and GFPf; lane d, GFPr and Neor; lane e, Neof and K9. (D) rKSHV.294 was tested for tTA control of SeAP expression by the infection of 293 cells or of 293 cells expressing tTA at an MOI of

    Techniques Used: Quantitation Assay, Construct, Polymerase Chain Reaction, Hybridization, Staining, Agarose Gel Electrophoresis, Electrophoresis, Expressing, Infection

    Related Articles

    Plasmid Preparation:

    Article Title: In vitro Enzymology of Cas9
    Article Snippet: .. For pUC19-based plasmids, use 50 units of SspI-HF (New England Biolabs) and 5 μg plasmid in 1 x CutSmart™ buffer in a total volume of 50 μl. ..

    Article Title: Computational Design of an Allosteric Antibody Switch by Deletion and Rescue of a Complex Structural Constellation
    Article Snippet: Plasmid DNA from liquid culture was isolated according to the protocol provided by the manufacturer (QIAprep Spin Miniprep Kit, QIAGEN). .. Next, the empty vector DNA was linearized using SspI-HF (NEB); the reaction was performed according to the protocol provided by the manufacturer, and the linearized vector was gel purified from a 0.8% agarose gel using a QIAquick gel extraction kit (QIAGEN). .. As for insert DNA, WT and mutant genes were synthesized in a double-stranded linear DNA fragment format (GeneArt Strings DNA Fragments, ThermoFisher).

    Southern Blot:

    Article Title: Nuclear pores as versatile reference standards for quantitative superresolution microscopy
    Article Snippet: .. Southern blotting was performed in accordance Koch et al. Genomic DNA was prepared using the Wizard Genomic DNA Purification kit (Promega) and digested with SspI-HF and MfeI-HF (NEB). ..

    DNA Purification:

    Article Title: Nuclear pores as versatile reference standards for quantitative superresolution microscopy
    Article Snippet: .. Southern blotting was performed in accordance Koch et al. Genomic DNA was prepared using the Wizard Genomic DNA Purification kit (Promega) and digested with SspI-HF and MfeI-HF (NEB). ..

    other:

    Article Title: Preparation of a New Construct of Human Histone Deacetylase 8 for the Crystallization of Enzyme-Inhibitor Complexes
    Article Snippet: 0.6 units/μL SspI-HF (3 μL of 20 units/μL stock).

    Purification:

    Article Title: Computational Design of an Allosteric Antibody Switch by Deletion and Rescue of a Complex Structural Constellation
    Article Snippet: Plasmid DNA from liquid culture was isolated according to the protocol provided by the manufacturer (QIAprep Spin Miniprep Kit, QIAGEN). .. Next, the empty vector DNA was linearized using SspI-HF (NEB); the reaction was performed according to the protocol provided by the manufacturer, and the linearized vector was gel purified from a 0.8% agarose gel using a QIAquick gel extraction kit (QIAGEN). .. As for insert DNA, WT and mutant genes were synthesized in a double-stranded linear DNA fragment format (GeneArt Strings DNA Fragments, ThermoFisher).

    Agarose Gel Electrophoresis:

    Article Title: Computational Design of an Allosteric Antibody Switch by Deletion and Rescue of a Complex Structural Constellation
    Article Snippet: Plasmid DNA from liquid culture was isolated according to the protocol provided by the manufacturer (QIAprep Spin Miniprep Kit, QIAGEN). .. Next, the empty vector DNA was linearized using SspI-HF (NEB); the reaction was performed according to the protocol provided by the manufacturer, and the linearized vector was gel purified from a 0.8% agarose gel using a QIAquick gel extraction kit (QIAGEN). .. As for insert DNA, WT and mutant genes were synthesized in a double-stranded linear DNA fragment format (GeneArt Strings DNA Fragments, ThermoFisher).

    Gel Extraction:

    Article Title: Computational Design of an Allosteric Antibody Switch by Deletion and Rescue of a Complex Structural Constellation
    Article Snippet: Plasmid DNA from liquid culture was isolated according to the protocol provided by the manufacturer (QIAprep Spin Miniprep Kit, QIAGEN). .. Next, the empty vector DNA was linearized using SspI-HF (NEB); the reaction was performed according to the protocol provided by the manufacturer, and the linearized vector was gel purified from a 0.8% agarose gel using a QIAquick gel extraction kit (QIAGEN). .. As for insert DNA, WT and mutant genes were synthesized in a double-stranded linear DNA fragment format (GeneArt Strings DNA Fragments, ThermoFisher).

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    New England Biolabs sspi hf
    H. pylori SS1 and PMSS1 are gene copy number variable at the cagA locus. (A) Schematic diagram showing tandem arrays of the identical 5,072-bp repeat regions at the cagA locus, all of which contain identical copies of the 3,540-bp cagA gene. The <t>SspI</t> fragment sizes of strains with 1 to 4 cagA copies are shown at the right. The drawing is not to scale. (B) Southern blot of SspI-digested genomic <t>DNA</t> from H. pylori SS1 or PMSS1 probed with a 297-bp PCR product amplified from SS1 cagA bp 1217 to 1514. The original working stocks of SS1 (lane 1) and PMSS1 (lane 5) showed bands corresponding in size to 4, 3, 2, and 1 copies of cagA (asterisks). Subculture of 4 single colonies from the freezer stocks demonstrated clones with 4, 3, or 2 copies of cagA (lanes 2 to 4) from SS1; subculture of 12 single colonies showed either 4 or 2 copies of cagA in PMSS1 (lanes 6 to 8). PMSS1 with a cagA deletion served as a negative control (lane 9). A kilobase ladder is shown at the left. (C) Western blot of H. pylori PMSS1 to examine whether the cagA copy number is positively correlated with protein expression. For this analysis, six individual single-colony isolates of PMSS1 were used, two each with four copies, two copies, or one copy of cagA . Relative quantities of protein in each band were determined using Image Lab software (Bio-Rad Laboratories). The density of the CagA band was divided by the density of the corresponding UreB band to obtain the normalized quantity of CagA to account for differences in gel loading.
    Sspi Hf, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sspi hf/product/New England Biolabs
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sspi hf - by Bioz Stars, 2021-03
    93/100 stars
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    H. pylori SS1 and PMSS1 are gene copy number variable at the cagA locus. (A) Schematic diagram showing tandem arrays of the identical 5,072-bp repeat regions at the cagA locus, all of which contain identical copies of the 3,540-bp cagA gene. The SspI fragment sizes of strains with 1 to 4 cagA copies are shown at the right. The drawing is not to scale. (B) Southern blot of SspI-digested genomic DNA from H. pylori SS1 or PMSS1 probed with a 297-bp PCR product amplified from SS1 cagA bp 1217 to 1514. The original working stocks of SS1 (lane 1) and PMSS1 (lane 5) showed bands corresponding in size to 4, 3, 2, and 1 copies of cagA (asterisks). Subculture of 4 single colonies from the freezer stocks demonstrated clones with 4, 3, or 2 copies of cagA (lanes 2 to 4) from SS1; subculture of 12 single colonies showed either 4 or 2 copies of cagA in PMSS1 (lanes 6 to 8). PMSS1 with a cagA deletion served as a negative control (lane 9). A kilobase ladder is shown at the left. (C) Western blot of H. pylori PMSS1 to examine whether the cagA copy number is positively correlated with protein expression. For this analysis, six individual single-colony isolates of PMSS1 were used, two each with four copies, two copies, or one copy of cagA . Relative quantities of protein in each band were determined using Image Lab software (Bio-Rad Laboratories). The density of the CagA band was divided by the density of the corresponding UreB band to obtain the normalized quantity of CagA to account for differences in gel loading.

    Journal: mBio

    Article Title: Fallacy of the Unique Genome: Sequence Diversity within Single Helicobacter pylori Strains

    doi: 10.1128/mBio.02321-16

    Figure Lengend Snippet: H. pylori SS1 and PMSS1 are gene copy number variable at the cagA locus. (A) Schematic diagram showing tandem arrays of the identical 5,072-bp repeat regions at the cagA locus, all of which contain identical copies of the 3,540-bp cagA gene. The SspI fragment sizes of strains with 1 to 4 cagA copies are shown at the right. The drawing is not to scale. (B) Southern blot of SspI-digested genomic DNA from H. pylori SS1 or PMSS1 probed with a 297-bp PCR product amplified from SS1 cagA bp 1217 to 1514. The original working stocks of SS1 (lane 1) and PMSS1 (lane 5) showed bands corresponding in size to 4, 3, 2, and 1 copies of cagA (asterisks). Subculture of 4 single colonies from the freezer stocks demonstrated clones with 4, 3, or 2 copies of cagA (lanes 2 to 4) from SS1; subculture of 12 single colonies showed either 4 or 2 copies of cagA in PMSS1 (lanes 6 to 8). PMSS1 with a cagA deletion served as a negative control (lane 9). A kilobase ladder is shown at the left. (C) Western blot of H. pylori PMSS1 to examine whether the cagA copy number is positively correlated with protein expression. For this analysis, six individual single-colony isolates of PMSS1 were used, two each with four copies, two copies, or one copy of cagA . Relative quantities of protein in each band were determined using Image Lab software (Bio-Rad Laboratories). The density of the CagA band was divided by the density of the corresponding UreB band to obtain the normalized quantity of CagA to account for differences in gel loading.

    Article Snippet: Genomic DNA was digested with SspI-HF (New England Biolabs) for 2 h. Digested DNA was separated on a 0.5% agarose gel overnight at 0.75 V/cm and transferred to a nylon membrane.

    Techniques: Southern Blot, Polymerase Chain Reaction, Amplification, Clone Assay, Negative Control, Western Blot, Expressing, Software

    Construction and quantitation of rKSHV.294. (A) Schematic diagram of rKSHV.294 showing the insertion site in the KSHV genome; the BamHI sites flanking the 4.8-kb segment of the KSHV genome used are indicated. The relative positions of the SeAP, the GFP, and the Neo elements are shown with their respective promoters but not to scale. Beneath the 294 construct are shown the expected PCR products (a to e) with the primers used for analysis of viral DNA. (B) Hybridization analysis of rKSHV.294 and JSC-1 viral DNA. (Left) KSHV probe using the 4.8-kb BamHI fragment used to construct the virus. (Right) Neo probe. Lane 1, rKSHV.294 × AflII, predicted fragments of 15.9, 5.7, and 4.8 kb. The 4.8-kb fragment contains the SeAP/GFP/Neo insert with only 170 bp of KSHV DNA, which accounts for the weak band. Lane 2, JSC-1 × AflII, predicted fragments of 15.9 and 5.8 kb. Lane 3, rKSHV.294 × SspI, predicted fragments of 8 and 6.2 kb. Lane 4, JSC-1 × SspI, predicted fragments of 6.2 and 3.3 kb. Lane 5, rKSHV.294 × AflII, predicted fragment of 4.8 kb. Lane 6, JSC-1 × AflII. Lane 7, rKSHV.294 DNA × SspI, predicted fragment of 8 kb. Lane 8, JSC-1 × SspI. DNA markers in kb are on the left side. (C) Ethidium bromide-stained agarose gel following electrophoresis of the PCR products resulting with rKSHV.294 viral DNA with the indicated primers described in Materials and Methods. Lane a, ORF56f (outside BamHI site) and SeAPr; lane b, ORF57u and SeAPr; lane c, SeAPf and GFPf; lane d, GFPr and Neor; lane e, Neof and K9. (D) rKSHV.294 was tested for tTA control of SeAP expression by the infection of 293 cells or of 293 cells expressing tTA at an MOI of

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: The HIV Protease Inhibitor Nelfinavir Inhibits Kaposi's Sarcoma-Associated Herpesvirus Replication In Vitro ▿

    doi: 10.1128/AAC.01295-10

    Figure Lengend Snippet: Construction and quantitation of rKSHV.294. (A) Schematic diagram of rKSHV.294 showing the insertion site in the KSHV genome; the BamHI sites flanking the 4.8-kb segment of the KSHV genome used are indicated. The relative positions of the SeAP, the GFP, and the Neo elements are shown with their respective promoters but not to scale. Beneath the 294 construct are shown the expected PCR products (a to e) with the primers used for analysis of viral DNA. (B) Hybridization analysis of rKSHV.294 and JSC-1 viral DNA. (Left) KSHV probe using the 4.8-kb BamHI fragment used to construct the virus. (Right) Neo probe. Lane 1, rKSHV.294 × AflII, predicted fragments of 15.9, 5.7, and 4.8 kb. The 4.8-kb fragment contains the SeAP/GFP/Neo insert with only 170 bp of KSHV DNA, which accounts for the weak band. Lane 2, JSC-1 × AflII, predicted fragments of 15.9 and 5.8 kb. Lane 3, rKSHV.294 × SspI, predicted fragments of 8 and 6.2 kb. Lane 4, JSC-1 × SspI, predicted fragments of 6.2 and 3.3 kb. Lane 5, rKSHV.294 × AflII, predicted fragment of 4.8 kb. Lane 6, JSC-1 × AflII. Lane 7, rKSHV.294 DNA × SspI, predicted fragment of 8 kb. Lane 8, JSC-1 × SspI. DNA markers in kb are on the left side. (C) Ethidium bromide-stained agarose gel following electrophoresis of the PCR products resulting with rKSHV.294 viral DNA with the indicated primers described in Materials and Methods. Lane a, ORF56f (outside BamHI site) and SeAPr; lane b, ORF57u and SeAPr; lane c, SeAPf and GFPf; lane d, GFPr and Neor; lane e, Neof and K9. (D) rKSHV.294 was tested for tTA control of SeAP expression by the infection of 293 cells or of 293 cells expressing tTA at an MOI of

    Article Snippet: Twenty micrograms of each DNA sample was digested overnight at 37°C with either AflII-HF or SspI-HF restriction endonucleases according to the manufacturer's recommendations (New England BioLabs, Ipswich, MA).

    Techniques: Quantitation Assay, Construct, Polymerase Chain Reaction, Hybridization, Staining, Agarose Gel Electrophoresis, Electrophoresis, Expressing, Infection