fsei  (New England Biolabs)


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    FseI
    Description:
    FseI 500 units
    Catalog Number:
    R0588L
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    Category:
    Restriction Enzymes
    Size:
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    New England Biolabs fsei
    FseI
    FseI 500 units
    https://www.bioz.com/result/fsei/product/New England Biolabs
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    fsei - by Bioz Stars, 2021-06
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    Images

    1) Product Images from "Extensive amplification of GI-VII-6, a multidrug resistance genomic island of Salmonella enterica serovar Typhimurium, increases resistance to extended-spectrum cephalosporins"

    Article Title: Extensive amplification of GI-VII-6, a multidrug resistance genomic island of Salmonella enterica serovar Typhimurium, increases resistance to extended-spectrum cephalosporins

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2015.00078

    PFGE–Southern hybridization images demonstrating bla CMY−2 and spvB locations in S . Typhimurium strains. (A) PFGE separation of S1 nuclease-digested genomic DNA from the selected strains followed by Southern hybridization with probes specific to bla CMY−2 and spvB . An arrow indicates the position of the endogenous 133-kb plasmid, in which the spvB signal was detected. Open triangles indicate the positions of enlarged plasmids, in which both bla CMY−2 and spvB signals were detected. Both signals were observed in multiple plasmid bands in strains 12-13, 12-14, and 12-18. (B) PFGE separation of FseI-digested genomic DNA from the selected strains followed by Southern hybridization with a bla CMY−2 probe. An arrow indicates the original position of the restricted fragment (350 kb) in which the chromosomal bla CMY−2 signal was detected. In the selected mutants, this fragment was not present and the bla CMY−2 signal was identified on the largest band as indicated by a closed trangle. Open triangles indicate the position of enlarged plasmid observed in strains 12-14, 25-6, and 25-17. (C) PFGE separation of XbaI-digested genomic DNA from the selected strains followed by Southern hybridization with a bla CMY−2 probe. An arrow indicates the original position of the restricted fragment (520 kb) in which chromosomal bla CMY−2 signal was detected. Closed triangles indicate the position of bla CMY−2 signals that originated from tandemly amplified GI-VII-6. (D) Restriction maps of the GI-VII-6 and its flanking region in the parent strain L-3553 and the mutants with putative amplifications. One putative amplified region presented is flanked by the first and the third copies of IS 26 (upper), and the other is flanked by the first and the fourth copies of IS 26 (lower). These amplified regions generate 100-kb (upper) and 125-kb (lower) fragments after XbaI digestion, respectively.
    Figure Legend Snippet: PFGE–Southern hybridization images demonstrating bla CMY−2 and spvB locations in S . Typhimurium strains. (A) PFGE separation of S1 nuclease-digested genomic DNA from the selected strains followed by Southern hybridization with probes specific to bla CMY−2 and spvB . An arrow indicates the position of the endogenous 133-kb plasmid, in which the spvB signal was detected. Open triangles indicate the positions of enlarged plasmids, in which both bla CMY−2 and spvB signals were detected. Both signals were observed in multiple plasmid bands in strains 12-13, 12-14, and 12-18. (B) PFGE separation of FseI-digested genomic DNA from the selected strains followed by Southern hybridization with a bla CMY−2 probe. An arrow indicates the original position of the restricted fragment (350 kb) in which the chromosomal bla CMY−2 signal was detected. In the selected mutants, this fragment was not present and the bla CMY−2 signal was identified on the largest band as indicated by a closed trangle. Open triangles indicate the position of enlarged plasmid observed in strains 12-14, 25-6, and 25-17. (C) PFGE separation of XbaI-digested genomic DNA from the selected strains followed by Southern hybridization with a bla CMY−2 probe. An arrow indicates the original position of the restricted fragment (520 kb) in which chromosomal bla CMY−2 signal was detected. Closed triangles indicate the position of bla CMY−2 signals that originated from tandemly amplified GI-VII-6. (D) Restriction maps of the GI-VII-6 and its flanking region in the parent strain L-3553 and the mutants with putative amplifications. One putative amplified region presented is flanked by the first and the third copies of IS 26 (upper), and the other is flanked by the first and the fourth copies of IS 26 (lower). These amplified regions generate 100-kb (upper) and 125-kb (lower) fragments after XbaI digestion, respectively.

    Techniques Used: Hybridization, Plasmid Preparation, Amplification

    2) Product Images from "senX3-independent contribution of regX3 to Mycobacterium tuberculosis virulence"

    Article Title: senX3-independent contribution of regX3 to Mycobacterium tuberculosis virulence

    Journal: BMC Microbiology

    doi: 10.1186/s12866-014-0265-8

    Genetic confirmation of complementation of senX3 ::Tn and regX3 ::Tn. (A) . Diagram of a 2824-bp fragment containing the senX3-regX3 operon from the genome of wild-type CDC1551. (B) FseI digestion of genomic DNA and hybridization with dig-labeled senX3 probe reveals fragments measuring 3.7-kb and 5.7-kb in CDC1551 and senX 3::Tn, respectively, as well as 5.7-kb and 7.7-kb fragments in senX3 ::Tn Comp. (C) SphI digestion of genomic DNA followed by hybridization with dig-labeled regX3 probe reveals 5.5-kb and 7.5-kb fragments in CDC1551 and regX3 ::Tn, respectively, as well as 7.5-kb and 4.6-kb fragments in regX3 ::Tn Comp. (D) Southern analysis of senX3 complementation. Lanes 1 and 2: SenX3 ::Tn Comp candidates with appropriately-sized bands; Lane 3: SenX 3::Tn; Lane 4: Wild-type CDC1551. (E) Southern analysis of regX3 complementation. Lane 1: Wild-type CDC551; Lane 2: regX 3::Tn; Lanes 3–5: regX3 ::Tn Comp candidates with appropriately-sized bands.
    Figure Legend Snippet: Genetic confirmation of complementation of senX3 ::Tn and regX3 ::Tn. (A) . Diagram of a 2824-bp fragment containing the senX3-regX3 operon from the genome of wild-type CDC1551. (B) FseI digestion of genomic DNA and hybridization with dig-labeled senX3 probe reveals fragments measuring 3.7-kb and 5.7-kb in CDC1551 and senX 3::Tn, respectively, as well as 5.7-kb and 7.7-kb fragments in senX3 ::Tn Comp. (C) SphI digestion of genomic DNA followed by hybridization with dig-labeled regX3 probe reveals 5.5-kb and 7.5-kb fragments in CDC1551 and regX3 ::Tn, respectively, as well as 7.5-kb and 4.6-kb fragments in regX3 ::Tn Comp. (D) Southern analysis of senX3 complementation. Lanes 1 and 2: SenX3 ::Tn Comp candidates with appropriately-sized bands; Lane 3: SenX 3::Tn; Lane 4: Wild-type CDC1551. (E) Southern analysis of regX3 complementation. Lane 1: Wild-type CDC551; Lane 2: regX 3::Tn; Lanes 3–5: regX3 ::Tn Comp candidates with appropriately-sized bands.

    Techniques Used: Hybridization, Labeling

    Related Articles

    Polymerase Chain Reaction:

    Article Title: Designing and Cloning Molecular Constructs to Knock Out N-Acetylglucosamine Phosphatidylinositol De-N-Acetylase (GPI12) Gene in Leishmania major (MRHO/IR/75/ER)
    Article Snippet: PLEXSY-neo2/R1 was extracted via the alkaline lysis method. .. The PCR fragments were cut by Fse I and Bsr gI enzymes (New England Biolabs, UK). .. The plasmid pGEM-B1 contains sequences of flank areas, which preform PCR by primer M13.

    Article Title: Structural Insights into the Catalytic Mechanism of a Plant Diterpene Glycosyltransferase SrUGT76G1
    Article Snippet: Subsequently, cDNA from A. thaliana was used as template and amplified with primer pair Ats3InfuYF09-Fse and Ats3InfuYF09-Kpn ( ) to give a fragment of AtSUS3 . .. The PCR product was then infused into Fse I and Kpn I (NEB) predigested vector pYF09, resulting in pLW108. .. To construct pHJ651 and pHJ830, we amplified the vector backbone from pLW108 with primer 830VF and 830VR, while insert fragment of SrUGT76G1 and SrUGT76G1_T284S was amplified from plasmid pQZ11 and pQZ11_T284S with primer pair inF and inR, respectively.

    Article Title: SIRT4 is essential for metabolic control and meiotic structure during mouse oocyte maturation, et al. SIRT4 is essential for metabolic control and meiotic structure during mouse oocyte maturation
    Article Snippet: 4.4 Plasmid construction and mRNA synthesis Total RNA was extracted from 100 mouse oocytes using Arcturus PicoPure RNA Isolation Kit (Applied Biosystems, CA, USA), and the cDNA was generated with QIAquick PCR Purification Kit (Qiagen, Germany). .. PCR products were purified, digested with FseI and AscI (NEB Inc, MA, USA), and then cloned into the pCS2+ vector with six Myc tags. .. The pCS2+ vectors encoding the PDHE1α and SIRT4H158Y mutants were generated with the use of a QuickChange site‐directed mutagenesis kit (Stratagene).

    Plasmid Preparation:

    Article Title: Construction of Gpm6a/ReelinGFPCreERT2 by BAC recombination using a specific gene in hepatic mesothelial or stellate cells
    Article Snippet: A single colony was picked and inoculated in LB medium with Kam and incubated for 4-6 h. The shuttle vector was purified with an Endofree plasmid maxi kit (Qiagen). .. The vector was digested with Not I and Fse I (New England Biolabs), and electrophoresis was performed with agarose gels (Takara, Osaka, Japan). .. The large fragment of GFPCreERT2 was extracted from the gel with a quick gel extraction kit (Qiagen).

    Article Title: Structural Insights into the Catalytic Mechanism of a Plant Diterpene Glycosyltransferase SrUGT76G1
    Article Snippet: Subsequently, cDNA from A. thaliana was used as template and amplified with primer pair Ats3InfuYF09-Fse and Ats3InfuYF09-Kpn ( ) to give a fragment of AtSUS3 . .. The PCR product was then infused into Fse I and Kpn I (NEB) predigested vector pYF09, resulting in pLW108. .. To construct pHJ651 and pHJ830, we amplified the vector backbone from pLW108 with primer 830VF and 830VR, while insert fragment of SrUGT76G1 and SrUGT76G1_T284S was amplified from plasmid pQZ11 and pQZ11_T284S with primer pair inF and inR, respectively.

    Article Title: Immunization with HIV-1 envelope T20-encoding DNA vaccines elicits cross-clade neutralizing antibody responses
    Article Snippet: One of these plasmids, pCMV-EnvB , was used as a donor vector for the expression cassette to produce pCMV-T20 and MC-T20 (see below). pCMV-DNA , which does not contain any expression cassette, was used for control immunizations. .. Minicircle MC-T20 ( ): The nucleotide (nt) sequence for the T20 peptide with a secretion signal (ss) from the V-J2-C region of the mouse Ig kappa-chain for efficient secretion of recombinant proteins (Immunomedics Inc., Morris Plains, NJ, USA) was cloned into the pCMV-T20 plasmid (see pCMV-EnvABC ) using Fse I and EcoRV restriction sites, by enzymes from New England Biolabs (R0588S and R0195S, NEB). .. PCR was performed to amplify the ss-T20 gene fragment (Eurofins Genomics, DE).

    Article Title: SIRT4 is essential for metabolic control and meiotic structure during mouse oocyte maturation, et al. SIRT4 is essential for metabolic control and meiotic structure during mouse oocyte maturation
    Article Snippet: 4.4 Plasmid construction and mRNA synthesis Total RNA was extracted from 100 mouse oocytes using Arcturus PicoPure RNA Isolation Kit (Applied Biosystems, CA, USA), and the cDNA was generated with QIAquick PCR Purification Kit (Qiagen, Germany). .. PCR products were purified, digested with FseI and AscI (NEB Inc, MA, USA), and then cloned into the pCS2+ vector with six Myc tags. .. The pCS2+ vectors encoding the PDHE1α and SIRT4H158Y mutants were generated with the use of a QuickChange site‐directed mutagenesis kit (Stratagene).

    Electrophoresis:

    Article Title: Construction of Gpm6a/ReelinGFPCreERT2 by BAC recombination using a specific gene in hepatic mesothelial or stellate cells
    Article Snippet: A single colony was picked and inoculated in LB medium with Kam and incubated for 4-6 h. The shuttle vector was purified with an Endofree plasmid maxi kit (Qiagen). .. The vector was digested with Not I and Fse I (New England Biolabs), and electrophoresis was performed with agarose gels (Takara, Osaka, Japan). .. The large fragment of GFPCreERT2 was extracted from the gel with a quick gel extraction kit (Qiagen).

    Article Title: senX3-independent contribution of regX3 to Mycobacterium tuberculosis virulence
    Article Snippet: In order to further confirm complementation of senX3 ::Tn and regX3 ::Tn, Southern blotting was performed using DIG High Prime DNA Labeling and Detection Starter Kit I according to the manufacturer’s protocol (Roche). .. Genomic DNA from senX3 ::Tn and regX3 ::Tn complement candidates was digested with FseI and SphI-HF (New England Biolabs), respectively, and electrophoresis was performed on 1% agarose gels. .. Following denaturation and neutralization, the gels were transferred onto positively charged nylon membranes (GE Healthcare).

    Sequencing:

    Article Title: Immunization with HIV-1 envelope T20-encoding DNA vaccines elicits cross-clade neutralizing antibody responses
    Article Snippet: One of these plasmids, pCMV-EnvB , was used as a donor vector for the expression cassette to produce pCMV-T20 and MC-T20 (see below). pCMV-DNA , which does not contain any expression cassette, was used for control immunizations. .. Minicircle MC-T20 ( ): The nucleotide (nt) sequence for the T20 peptide with a secretion signal (ss) from the V-J2-C region of the mouse Ig kappa-chain for efficient secretion of recombinant proteins (Immunomedics Inc., Morris Plains, NJ, USA) was cloned into the pCMV-T20 plasmid (see pCMV-EnvABC ) using Fse I and EcoRV restriction sites, by enzymes from New England Biolabs (R0588S and R0195S, NEB). .. PCR was performed to amplify the ss-T20 gene fragment (Eurofins Genomics, DE).

    Recombinant:

    Article Title: Immunization with HIV-1 envelope T20-encoding DNA vaccines elicits cross-clade neutralizing antibody responses
    Article Snippet: One of these plasmids, pCMV-EnvB , was used as a donor vector for the expression cassette to produce pCMV-T20 and MC-T20 (see below). pCMV-DNA , which does not contain any expression cassette, was used for control immunizations. .. Minicircle MC-T20 ( ): The nucleotide (nt) sequence for the T20 peptide with a secretion signal (ss) from the V-J2-C region of the mouse Ig kappa-chain for efficient secretion of recombinant proteins (Immunomedics Inc., Morris Plains, NJ, USA) was cloned into the pCMV-T20 plasmid (see pCMV-EnvABC ) using Fse I and EcoRV restriction sites, by enzymes from New England Biolabs (R0588S and R0195S, NEB). .. PCR was performed to amplify the ss-T20 gene fragment (Eurofins Genomics, DE).

    Clone Assay:

    Article Title: Immunization with HIV-1 envelope T20-encoding DNA vaccines elicits cross-clade neutralizing antibody responses
    Article Snippet: One of these plasmids, pCMV-EnvB , was used as a donor vector for the expression cassette to produce pCMV-T20 and MC-T20 (see below). pCMV-DNA , which does not contain any expression cassette, was used for control immunizations. .. Minicircle MC-T20 ( ): The nucleotide (nt) sequence for the T20 peptide with a secretion signal (ss) from the V-J2-C region of the mouse Ig kappa-chain for efficient secretion of recombinant proteins (Immunomedics Inc., Morris Plains, NJ, USA) was cloned into the pCMV-T20 plasmid (see pCMV-EnvABC ) using Fse I and EcoRV restriction sites, by enzymes from New England Biolabs (R0588S and R0195S, NEB). .. PCR was performed to amplify the ss-T20 gene fragment (Eurofins Genomics, DE).

    Article Title: SIRT4 is essential for metabolic control and meiotic structure during mouse oocyte maturation, et al. SIRT4 is essential for metabolic control and meiotic structure during mouse oocyte maturation
    Article Snippet: 4.4 Plasmid construction and mRNA synthesis Total RNA was extracted from 100 mouse oocytes using Arcturus PicoPure RNA Isolation Kit (Applied Biosystems, CA, USA), and the cDNA was generated with QIAquick PCR Purification Kit (Qiagen, Germany). .. PCR products were purified, digested with FseI and AscI (NEB Inc, MA, USA), and then cloned into the pCS2+ vector with six Myc tags. .. The pCS2+ vectors encoding the PDHE1α and SIRT4H158Y mutants were generated with the use of a QuickChange site‐directed mutagenesis kit (Stratagene).

    Purification:

    Article Title: SIRT4 is essential for metabolic control and meiotic structure during mouse oocyte maturation, et al. SIRT4 is essential for metabolic control and meiotic structure during mouse oocyte maturation
    Article Snippet: 4.4 Plasmid construction and mRNA synthesis Total RNA was extracted from 100 mouse oocytes using Arcturus PicoPure RNA Isolation Kit (Applied Biosystems, CA, USA), and the cDNA was generated with QIAquick PCR Purification Kit (Qiagen, Germany). .. PCR products were purified, digested with FseI and AscI (NEB Inc, MA, USA), and then cloned into the pCS2+ vector with six Myc tags. .. The pCS2+ vectors encoding the PDHE1α and SIRT4H158Y mutants were generated with the use of a QuickChange site‐directed mutagenesis kit (Stratagene).

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    New England Biolabs fsei
    PFGE–Southern hybridization images demonstrating bla CMY−2 and spvB locations in S . Typhimurium strains. (A) PFGE separation of S1 nuclease-digested genomic DNA from the selected strains followed by Southern hybridization with probes specific to bla CMY−2 and spvB . An arrow indicates the position of the endogenous 133-kb plasmid, in which the spvB signal was detected. Open triangles indicate the positions of enlarged plasmids, in which both bla CMY−2 and spvB signals were detected. Both signals were observed in multiple plasmid bands in strains 12-13, 12-14, and 12-18. (B) PFGE separation of <t>FseI-digested</t> genomic DNA from the selected strains followed by Southern hybridization with a bla CMY−2 probe. An arrow indicates the original position of the restricted fragment (350 kb) in which the chromosomal bla CMY−2 signal was detected. In the selected mutants, this fragment was not present and the bla CMY−2 signal was identified on the largest band as indicated by a closed trangle. Open triangles indicate the position of enlarged plasmid observed in strains 12-14, 25-6, and 25-17. (C) PFGE separation of <t>XbaI-digested</t> genomic DNA from the selected strains followed by Southern hybridization with a bla CMY−2 probe. An arrow indicates the original position of the restricted fragment (520 kb) in which chromosomal bla CMY−2 signal was detected. Closed triangles indicate the position of bla CMY−2 signals that originated from tandemly amplified GI-VII-6. (D) Restriction maps of the GI-VII-6 and its flanking region in the parent strain L-3553 and the mutants with putative amplifications. One putative amplified region presented is flanked by the first and the third copies of IS 26 (upper), and the other is flanked by the first and the fourth copies of IS 26 (lower). These amplified regions generate 100-kb (upper) and 125-kb (lower) fragments after XbaI digestion, respectively.
    Fsei, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    PFGE–Southern hybridization images demonstrating bla CMY−2 and spvB locations in S . Typhimurium strains. (A) PFGE separation of S1 nuclease-digested genomic DNA from the selected strains followed by Southern hybridization with probes specific to bla CMY−2 and spvB . An arrow indicates the position of the endogenous 133-kb plasmid, in which the spvB signal was detected. Open triangles indicate the positions of enlarged plasmids, in which both bla CMY−2 and spvB signals were detected. Both signals were observed in multiple plasmid bands in strains 12-13, 12-14, and 12-18. (B) PFGE separation of FseI-digested genomic DNA from the selected strains followed by Southern hybridization with a bla CMY−2 probe. An arrow indicates the original position of the restricted fragment (350 kb) in which the chromosomal bla CMY−2 signal was detected. In the selected mutants, this fragment was not present and the bla CMY−2 signal was identified on the largest band as indicated by a closed trangle. Open triangles indicate the position of enlarged plasmid observed in strains 12-14, 25-6, and 25-17. (C) PFGE separation of XbaI-digested genomic DNA from the selected strains followed by Southern hybridization with a bla CMY−2 probe. An arrow indicates the original position of the restricted fragment (520 kb) in which chromosomal bla CMY−2 signal was detected. Closed triangles indicate the position of bla CMY−2 signals that originated from tandemly amplified GI-VII-6. (D) Restriction maps of the GI-VII-6 and its flanking region in the parent strain L-3553 and the mutants with putative amplifications. One putative amplified region presented is flanked by the first and the third copies of IS 26 (upper), and the other is flanked by the first and the fourth copies of IS 26 (lower). These amplified regions generate 100-kb (upper) and 125-kb (lower) fragments after XbaI digestion, respectively.

    Journal: Frontiers in Microbiology

    Article Title: Extensive amplification of GI-VII-6, a multidrug resistance genomic island of Salmonella enterica serovar Typhimurium, increases resistance to extended-spectrum cephalosporins

    doi: 10.3389/fmicb.2015.00078

    Figure Lengend Snippet: PFGE–Southern hybridization images demonstrating bla CMY−2 and spvB locations in S . Typhimurium strains. (A) PFGE separation of S1 nuclease-digested genomic DNA from the selected strains followed by Southern hybridization with probes specific to bla CMY−2 and spvB . An arrow indicates the position of the endogenous 133-kb plasmid, in which the spvB signal was detected. Open triangles indicate the positions of enlarged plasmids, in which both bla CMY−2 and spvB signals were detected. Both signals were observed in multiple plasmid bands in strains 12-13, 12-14, and 12-18. (B) PFGE separation of FseI-digested genomic DNA from the selected strains followed by Southern hybridization with a bla CMY−2 probe. An arrow indicates the original position of the restricted fragment (350 kb) in which the chromosomal bla CMY−2 signal was detected. In the selected mutants, this fragment was not present and the bla CMY−2 signal was identified on the largest band as indicated by a closed trangle. Open triangles indicate the position of enlarged plasmid observed in strains 12-14, 25-6, and 25-17. (C) PFGE separation of XbaI-digested genomic DNA from the selected strains followed by Southern hybridization with a bla CMY−2 probe. An arrow indicates the original position of the restricted fragment (520 kb) in which chromosomal bla CMY−2 signal was detected. Closed triangles indicate the position of bla CMY−2 signals that originated from tandemly amplified GI-VII-6. (D) Restriction maps of the GI-VII-6 and its flanking region in the parent strain L-3553 and the mutants with putative amplifications. One putative amplified region presented is flanked by the first and the third copies of IS 26 (upper), and the other is flanked by the first and the fourth copies of IS 26 (lower). These amplified regions generate 100-kb (upper) and 125-kb (lower) fragments after XbaI digestion, respectively.

    Article Snippet: The plugs were digested with 60 U/ml of XbaI (Takara Bio Inc.) or 40 U/ml of FseI (New England BioLabs, Ipswich, MA, USA) at 37°C for 6 h, or 2 U/ml of S1 nuclease (Takara Bio Inc.) at 37°C for 45 min. Primers used for probe synthesis were listed in Table .

    Techniques: Hybridization, Plasmid Preparation, Amplification

    (A) The flank area with enzymes sequences (yellow: Bgl II, gray: Swa I, pink: Kpn I, orange: Fse I, violet: Bsr gI). (B) The map of the pGEM-b1 vector. (C) pGEM-b1 vector that contain the flank regions and specific restricted enzymes for cloning. (D) The pLexsy-2 vector contain construct after cloning

    Journal: Iranian Journal of Parasitology

    Article Title: Designing and Cloning Molecular Constructs to Knock Out N-Acetylglucosamine Phosphatidylinositol De-N-Acetylase (GPI12) Gene in Leishmania major (MRHO/IR/75/ER)

    doi:

    Figure Lengend Snippet: (A) The flank area with enzymes sequences (yellow: Bgl II, gray: Swa I, pink: Kpn I, orange: Fse I, violet: Bsr gI). (B) The map of the pGEM-b1 vector. (C) pGEM-b1 vector that contain the flank regions and specific restricted enzymes for cloning. (D) The pLexsy-2 vector contain construct after cloning

    Article Snippet: The PCR fragments were cut by Fse I and Bsr gI enzymes (New England Biolabs, UK).

    Techniques: Plasmid Preparation, Clone Assay, Construct

    In all figures, lane 1 shows molecular size marker. (A) The sequences of flank areas on which PCR was performed by primerM13; Lane 2 shows the amplified ∼1kb band. (B) The PCR fragments which were cut with Bgl II and Kpn I enzymes. The PCR product which was divided into 3 pieces of 600, 300 and 100 bp; Lane 2 shows the 300 bp fragment. (C) The pLEXSY vector, which was cut by Fse I and Bsr gI enzymes; Lane 2 shows the ∼ 0.7 kb fragment is separated from the vector. (D) The pLEXSY vector, which was cut by Bgl II and Kpn I enzymes. Lane 2 shows the ∼1 kb fragment is separated from the vector. (E) The PCR fragments which were cut with Fse I and Bsr gI enzymes. The PCR product which was divided into 3 pieces of 500, 400 and 100 bp; Lane 2 shows the 500 bp fragment

    Journal: Iranian Journal of Parasitology

    Article Title: Designing and Cloning Molecular Constructs to Knock Out N-Acetylglucosamine Phosphatidylinositol De-N-Acetylase (GPI12) Gene in Leishmania major (MRHO/IR/75/ER)

    doi:

    Figure Lengend Snippet: In all figures, lane 1 shows molecular size marker. (A) The sequences of flank areas on which PCR was performed by primerM13; Lane 2 shows the amplified ∼1kb band. (B) The PCR fragments which were cut with Bgl II and Kpn I enzymes. The PCR product which was divided into 3 pieces of 600, 300 and 100 bp; Lane 2 shows the 300 bp fragment. (C) The pLEXSY vector, which was cut by Fse I and Bsr gI enzymes; Lane 2 shows the ∼ 0.7 kb fragment is separated from the vector. (D) The pLEXSY vector, which was cut by Bgl II and Kpn I enzymes. Lane 2 shows the ∼1 kb fragment is separated from the vector. (E) The PCR fragments which were cut with Fse I and Bsr gI enzymes. The PCR product which was divided into 3 pieces of 500, 400 and 100 bp; Lane 2 shows the 500 bp fragment

    Article Snippet: The PCR fragments were cut by Fse I and Bsr gI enzymes (New England Biolabs, UK).

    Techniques: Marker, Polymerase Chain Reaction, Amplification, Plasmid Preparation

    Preparation of the reporter gene GFPCreERT2. A: Diagram of the shuttle vector carrying GFPCreERT2; B: Electrophoretogram of the shuttle vector cut by Not I and Fse I; C: Electrophoretogram after the GFPCreERT2 fragment was cut from the gel. M: Marker; Gpm6a: Shuttle vector inserted by the 5arm and 3arm from the Gpm6a BAC; Reelin: Shuttle vector inserted by the 5arm and 3arm from the Reelin BAC.

    Journal: World Journal of Gastroenterology

    Article Title: Construction of Gpm6a/ReelinGFPCreERT2 by BAC recombination using a specific gene in hepatic mesothelial or stellate cells

    doi: 10.3748/wjg.v23.i2.224

    Figure Lengend Snippet: Preparation of the reporter gene GFPCreERT2. A: Diagram of the shuttle vector carrying GFPCreERT2; B: Electrophoretogram of the shuttle vector cut by Not I and Fse I; C: Electrophoretogram after the GFPCreERT2 fragment was cut from the gel. M: Marker; Gpm6a: Shuttle vector inserted by the 5arm and 3arm from the Gpm6a BAC; Reelin: Shuttle vector inserted by the 5arm and 3arm from the Reelin BAC.

    Article Snippet: The vector was digested with Not I and Fse I (New England Biolabs), and electrophoresis was performed with agarose gels (Takara, Osaka, Japan).

    Techniques: Plasmid Preparation, Marker, BAC Assay

    Genetic confirmation of complementation of senX3 ::Tn and regX3 ::Tn. (A) . Diagram of a 2824-bp fragment containing the senX3-regX3 operon from the genome of wild-type CDC1551. (B) FseI digestion of genomic DNA and hybridization with dig-labeled senX3 probe reveals fragments measuring 3.7-kb and 5.7-kb in CDC1551 and senX 3::Tn, respectively, as well as 5.7-kb and 7.7-kb fragments in senX3 ::Tn Comp. (C) SphI digestion of genomic DNA followed by hybridization with dig-labeled regX3 probe reveals 5.5-kb and 7.5-kb fragments in CDC1551 and regX3 ::Tn, respectively, as well as 7.5-kb and 4.6-kb fragments in regX3 ::Tn Comp. (D) Southern analysis of senX3 complementation. Lanes 1 and 2: SenX3 ::Tn Comp candidates with appropriately-sized bands; Lane 3: SenX 3::Tn; Lane 4: Wild-type CDC1551. (E) Southern analysis of regX3 complementation. Lane 1: Wild-type CDC551; Lane 2: regX 3::Tn; Lanes 3–5: regX3 ::Tn Comp candidates with appropriately-sized bands.

    Journal: BMC Microbiology

    Article Title: senX3-independent contribution of regX3 to Mycobacterium tuberculosis virulence

    doi: 10.1186/s12866-014-0265-8

    Figure Lengend Snippet: Genetic confirmation of complementation of senX3 ::Tn and regX3 ::Tn. (A) . Diagram of a 2824-bp fragment containing the senX3-regX3 operon from the genome of wild-type CDC1551. (B) FseI digestion of genomic DNA and hybridization with dig-labeled senX3 probe reveals fragments measuring 3.7-kb and 5.7-kb in CDC1551 and senX 3::Tn, respectively, as well as 5.7-kb and 7.7-kb fragments in senX3 ::Tn Comp. (C) SphI digestion of genomic DNA followed by hybridization with dig-labeled regX3 probe reveals 5.5-kb and 7.5-kb fragments in CDC1551 and regX3 ::Tn, respectively, as well as 7.5-kb and 4.6-kb fragments in regX3 ::Tn Comp. (D) Southern analysis of senX3 complementation. Lanes 1 and 2: SenX3 ::Tn Comp candidates with appropriately-sized bands; Lane 3: SenX 3::Tn; Lane 4: Wild-type CDC1551. (E) Southern analysis of regX3 complementation. Lane 1: Wild-type CDC551; Lane 2: regX 3::Tn; Lanes 3–5: regX3 ::Tn Comp candidates with appropriately-sized bands.

    Article Snippet: Genomic DNA from senX3 ::Tn and regX3 ::Tn complement candidates was digested with FseI and SphI-HF (New England Biolabs), respectively, and electrophoresis was performed on 1% agarose gels.

    Techniques: Hybridization, Labeling