c2925  (New England Biolabs)


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    Name:
    dam dcm Competent E coli
    Description:
    dam dcm Competent E coli 20x0 05 ml
    Catalog Number:
    c2925h
    Price:
    246
    Size:
    1 ml
    Category:
    Competent Bacteria
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    New England Biolabs c2925
    dam dcm Competent E coli
    dam dcm Competent E coli 20x0 05 ml
    https://www.bioz.com/result/c2925/product/New England Biolabs
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    c2925 - by Bioz Stars, 2020-05
    95/100 stars

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    Related Articles

    Acetylene Reduction Assay:

    Article Title: Rapid targeted gene disruption in Bacillus anthracis
    Article Snippet: .. Since transformation efficiency of Bacillus anthracis is sensitive to dam methylation [ ], plasmid DNA for transformation into Bacillus anthracis was prepared from a dam methylase deficient host E. coli (New England Biolabs C2925H) with genotype: ara-14 leuB6 fhuA31 lacY1 tsx78 glnV44 galK2 galT22 mcrA dcm-6 hisG4 rfbD1 R(zgb210::Tn10) TetS endA1 rspL136 (StrR ) dam13::Tn9 (CamR ) xylA-5 mtl-1 thi-1 mcrB1 hsdR2. .. Bacillus anthracis (Sterne 34 F2) was obtained from Colorado Serum Co. (Denver, CO).

    Modification:

    Article Title: Target-dependent nickase activities of CRISPR-Cas nucleases Cpf1 and Cas9
    Article Snippet: .. Recombinant proteins were expressed in modified E. coli NiCo21 (DE3) cells (NEB C2925H) harboring the Cpf1 expression plasmid by growing in LB at 23°C for 16 hr in presence of IPTG at 0.4mM. .. As/Fn/Lb-Cpf1 was purified using HiTrap DEAE FF (GE Healthcare), HisTrap HP (Ni-NTA) (GE Healthcare) and HiTrapSP HP (GE Healthcare) columns.

    Methylation:

    Article Title: Metabolic engineering of Zymomonas mobilis for 2,3-butanediol production from lignocellulosic biomass sugars
    Article Snippet: .. Due to the presence of restriction/modification systems in Z. mobilis [ ] that can decrease transformation efficiency, all plasmids were built in and isolated from a methylation-deficient E. coli strain, C2925 (NEB, MA), for efficient transformation into Z. mobilis 8b or its derivatives. .. For single colony isolation, transformants grown on the selective plates containing spectinomycin were further streaked on RMG plates with spectinomycin at a final concentration of 200 μg/mL (RMGSp).

    Article Title: Rapid targeted gene disruption in Bacillus anthracis
    Article Snippet: .. Since transformation efficiency of Bacillus anthracis is sensitive to dam methylation [ ], plasmid DNA for transformation into Bacillus anthracis was prepared from a dam methylase deficient host E. coli (New England Biolabs C2925H) with genotype: ara-14 leuB6 fhuA31 lacY1 tsx78 glnV44 galK2 galT22 mcrA dcm-6 hisG4 rfbD1 R(zgb210::Tn10) TetS endA1 rspL136 (StrR ) dam13::Tn9 (CamR ) xylA-5 mtl-1 thi-1 mcrB1 hsdR2. .. Bacillus anthracis (Sterne 34 F2) was obtained from Colorado Serum Co. (Denver, CO).

    Isolation:

    Article Title: Metabolic engineering of Zymomonas mobilis for 2,3-butanediol production from lignocellulosic biomass sugars
    Article Snippet: .. Due to the presence of restriction/modification systems in Z. mobilis [ ] that can decrease transformation efficiency, all plasmids were built in and isolated from a methylation-deficient E. coli strain, C2925 (NEB, MA), for efficient transformation into Z. mobilis 8b or its derivatives. .. For single colony isolation, transformants grown on the selective plates containing spectinomycin were further streaked on RMG plates with spectinomycin at a final concentration of 200 μg/mL (RMGSp).

    Ligation:

    Article Title: The ATRX cDNA is prone to bacterial IS10 element insertions that alter its structure
    Article Snippet: .. Ligation product was transformed into dam- /dcm- competent E. coli (NEB C2925H). ..

    Article Title: Chlamydia trachomatis transformation and allelic exchange mutagenesis
    Article Snippet: .. pSUmC plasmid DNA pUC18A plasmid DNA pUC19G plasmid DNA Primer pair to amplify the gene targeted for deletion surrounded by approximately 3-kilobase (kb) flanking sequences from chlamydial genomic DNA: X@pUC F and R Primer pair surrounding, but directed away from, the target gene: X-away F and R Primer pair for amplifying the bla-gfp cassette from pUC19G: bla-gfp F: GGAAATGTGCGCGGAACCC bla-gfp R: TTACTTGTATAGTTCATCCATGCCATGTG Primer pair for amplifying the homologous recombination sequence for insertion into pSUmC: HomRR@pSUmC F: CTGCAGGTACCGGTCGACCATTC GGTCTGACGCTCAGTGGAACG HomRR@pSUmC R: GATCTTTCTACGGGGTCTGACGCTC CTGGCGTTACCCAACTTAATCGCC Q5® High-Fidelity DNA Polymerase (M0491S, NEB) DpnI restriction enzyme (R0176S, NEB) Cutsmart buffer (B7204S, NEB) T4 Polynucleotide Kinase (M0201S, NEB) Quick Ligation™ Kit (M2200S, NEB) NEB® 10-beta Competent E. coli (C3019H, NEB) dam− /dcm− Competent E. coli (C2925I, NEB) Zyppy™ Plasmid Miniprep Kit (D4036, Zymo Research) Zymoclean™ Gel DNA Recovery Kit (D4001, Zymo Research) QIAfilter Plasmid Maxi Kit (12262, Qiagen) Spectinomycin dihydrochloride pentahydrate (J61820, Alfa Aesar) Carbenicillin disodium salt (J61949, Alfa Aesar) LB Media (J106, VWR) Agar Powder ( , Alfa Aesar) Agarose LE (BMK-A1705, KSE Scientific) Phenol–Chloroform (6805, EMD Millipore) 3M sodium acetate 100% ethanol Design primers for amplifying gene × flanked upstream and downstream (±) by 3 kb arms for insertion into pUC18A. .. Sequences should be designed with melting temperatures of approximately 60°C which hairpin at temperatures no higher than 40°C, as determined by OligoAnalyzer 3.1 (Integrated DNA Technologies).

    Sequencing:

    Article Title: Chlamydia trachomatis transformation and allelic exchange mutagenesis
    Article Snippet: .. pSUmC plasmid DNA pUC18A plasmid DNA pUC19G plasmid DNA Primer pair to amplify the gene targeted for deletion surrounded by approximately 3-kilobase (kb) flanking sequences from chlamydial genomic DNA: X@pUC F and R Primer pair surrounding, but directed away from, the target gene: X-away F and R Primer pair for amplifying the bla-gfp cassette from pUC19G: bla-gfp F: GGAAATGTGCGCGGAACCC bla-gfp R: TTACTTGTATAGTTCATCCATGCCATGTG Primer pair for amplifying the homologous recombination sequence for insertion into pSUmC: HomRR@pSUmC F: CTGCAGGTACCGGTCGACCATTC GGTCTGACGCTCAGTGGAACG HomRR@pSUmC R: GATCTTTCTACGGGGTCTGACGCTC CTGGCGTTACCCAACTTAATCGCC Q5® High-Fidelity DNA Polymerase (M0491S, NEB) DpnI restriction enzyme (R0176S, NEB) Cutsmart buffer (B7204S, NEB) T4 Polynucleotide Kinase (M0201S, NEB) Quick Ligation™ Kit (M2200S, NEB) NEB® 10-beta Competent E. coli (C3019H, NEB) dam− /dcm− Competent E. coli (C2925I, NEB) Zyppy™ Plasmid Miniprep Kit (D4036, Zymo Research) Zymoclean™ Gel DNA Recovery Kit (D4001, Zymo Research) QIAfilter Plasmid Maxi Kit (12262, Qiagen) Spectinomycin dihydrochloride pentahydrate (J61820, Alfa Aesar) Carbenicillin disodium salt (J61949, Alfa Aesar) LB Media (J106, VWR) Agar Powder ( , Alfa Aesar) Agarose LE (BMK-A1705, KSE Scientific) Phenol–Chloroform (6805, EMD Millipore) 3M sodium acetate 100% ethanol Design primers for amplifying gene × flanked upstream and downstream (±) by 3 kb arms for insertion into pUC18A. .. Sequences should be designed with melting temperatures of approximately 60°C which hairpin at temperatures no higher than 40°C, as determined by OligoAnalyzer 3.1 (Integrated DNA Technologies).

    other:

    Article Title: Sources of artifact in measurements of 6mA and 4mC abundance in eukaryotic genomic DNA
    Article Snippet: Worms were grown on dam − dcm − bacteria (NEB C2925) on standard NGM plates in all experiments.

    Plasmid Preparation:

    Article Title: Target-dependent nickase activities of CRISPR-Cas nucleases Cpf1 and Cas9
    Article Snippet: .. Recombinant proteins were expressed in modified E. coli NiCo21 (DE3) cells (NEB C2925H) harboring the Cpf1 expression plasmid by growing in LB at 23°C for 16 hr in presence of IPTG at 0.4mM. .. As/Fn/Lb-Cpf1 was purified using HiTrap DEAE FF (GE Healthcare), HisTrap HP (Ni-NTA) (GE Healthcare) and HiTrapSP HP (GE Healthcare) columns.

    Article Title: Chlamydia trachomatis transformation and allelic exchange mutagenesis
    Article Snippet: .. pSUmC plasmid DNA pUC18A plasmid DNA pUC19G plasmid DNA Primer pair to amplify the gene targeted for deletion surrounded by approximately 3-kilobase (kb) flanking sequences from chlamydial genomic DNA: X@pUC F and R Primer pair surrounding, but directed away from, the target gene: X-away F and R Primer pair for amplifying the bla-gfp cassette from pUC19G: bla-gfp F: GGAAATGTGCGCGGAACCC bla-gfp R: TTACTTGTATAGTTCATCCATGCCATGTG Primer pair for amplifying the homologous recombination sequence for insertion into pSUmC: HomRR@pSUmC F: CTGCAGGTACCGGTCGACCATTC GGTCTGACGCTCAGTGGAACG HomRR@pSUmC R: GATCTTTCTACGGGGTCTGACGCTC CTGGCGTTACCCAACTTAATCGCC Q5® High-Fidelity DNA Polymerase (M0491S, NEB) DpnI restriction enzyme (R0176S, NEB) Cutsmart buffer (B7204S, NEB) T4 Polynucleotide Kinase (M0201S, NEB) Quick Ligation™ Kit (M2200S, NEB) NEB® 10-beta Competent E. coli (C3019H, NEB) dam− /dcm− Competent E. coli (C2925I, NEB) Zyppy™ Plasmid Miniprep Kit (D4036, Zymo Research) Zymoclean™ Gel DNA Recovery Kit (D4001, Zymo Research) QIAfilter Plasmid Maxi Kit (12262, Qiagen) Spectinomycin dihydrochloride pentahydrate (J61820, Alfa Aesar) Carbenicillin disodium salt (J61949, Alfa Aesar) LB Media (J106, VWR) Agar Powder ( , Alfa Aesar) Agarose LE (BMK-A1705, KSE Scientific) Phenol–Chloroform (6805, EMD Millipore) 3M sodium acetate 100% ethanol Design primers for amplifying gene × flanked upstream and downstream (±) by 3 kb arms for insertion into pUC18A. .. Sequences should be designed with melting temperatures of approximately 60°C which hairpin at temperatures no higher than 40°C, as determined by OligoAnalyzer 3.1 (Integrated DNA Technologies).

    Article Title: Rapid targeted gene disruption in Bacillus anthracis
    Article Snippet: .. Since transformation efficiency of Bacillus anthracis is sensitive to dam methylation [ ], plasmid DNA for transformation into Bacillus anthracis was prepared from a dam methylase deficient host E. coli (New England Biolabs C2925H) with genotype: ara-14 leuB6 fhuA31 lacY1 tsx78 glnV44 galK2 galT22 mcrA dcm-6 hisG4 rfbD1 R(zgb210::Tn10) TetS endA1 rspL136 (StrR ) dam13::Tn9 (CamR ) xylA-5 mtl-1 thi-1 mcrB1 hsdR2. .. Bacillus anthracis (Sterne 34 F2) was obtained from Colorado Serum Co. (Denver, CO).

    Expressing:

    Article Title: Target-dependent nickase activities of CRISPR-Cas nucleases Cpf1 and Cas9
    Article Snippet: .. Recombinant proteins were expressed in modified E. coli NiCo21 (DE3) cells (NEB C2925H) harboring the Cpf1 expression plasmid by growing in LB at 23°C for 16 hr in presence of IPTG at 0.4mM. .. As/Fn/Lb-Cpf1 was purified using HiTrap DEAE FF (GE Healthcare), HisTrap HP (Ni-NTA) (GE Healthcare) and HiTrapSP HP (GE Healthcare) columns.

    Polymerase Chain Reaction:

    Article Title: Transcriptomic Profiles of Zymomonas mobilis 8b to Furfural Acute and Long-Term Stress in Both Glucose and Xylose Conditions
    Article Snippet: .. NEB C2925 competent cells (NEB, Ipswich, MA, United States) were used for E. coli transformation, and transformants were confirmed by colony PCR using the primers of 130_SF/SR. .. The gene-specific primers were used to confirm that the targeted genes were cloned into the vector.

    Transformation Assay:

    Article Title: Modeling Cystic Fibrosis Using Pluripotent Stem Cell-Derived Human Pancreatic Ductal Epithelial Cells
    Article Snippet: .. The two products were then ligated and transformed in Dam− /Dcm− competent Escherichia coli (New England Biolabs, Ipswich, MA, ). .. The minimum CFTR promoter (372 bp) was inserted into the newly created plasmid by digestion of both the plasmid and the PCR product with EcoRI and AgeI.

    Article Title: The ATRX cDNA is prone to bacterial IS10 element insertions that alter its structure
    Article Snippet: .. Ligation product was transformed into dam- /dcm- competent E. coli (NEB C2925H). ..

    Article Title: Metabolic engineering of Zymomonas mobilis for 2,3-butanediol production from lignocellulosic biomass sugars
    Article Snippet: .. Due to the presence of restriction/modification systems in Z. mobilis [ ] that can decrease transformation efficiency, all plasmids were built in and isolated from a methylation-deficient E. coli strain, C2925 (NEB, MA), for efficient transformation into Z. mobilis 8b or its derivatives. .. For single colony isolation, transformants grown on the selective plates containing spectinomycin were further streaked on RMG plates with spectinomycin at a final concentration of 200 μg/mL (RMGSp).

    Article Title: Transcriptomic Profiles of Zymomonas mobilis 8b to Furfural Acute and Long-Term Stress in Both Glucose and Xylose Conditions
    Article Snippet: .. NEB C2925 competent cells (NEB, Ipswich, MA, United States) were used for E. coli transformation, and transformants were confirmed by colony PCR using the primers of 130_SF/SR. .. The gene-specific primers were used to confirm that the targeted genes were cloned into the vector.

    Article Title: Rapid targeted gene disruption in Bacillus anthracis
    Article Snippet: .. Since transformation efficiency of Bacillus anthracis is sensitive to dam methylation [ ], plasmid DNA for transformation into Bacillus anthracis was prepared from a dam methylase deficient host E. coli (New England Biolabs C2925H) with genotype: ara-14 leuB6 fhuA31 lacY1 tsx78 glnV44 galK2 galT22 mcrA dcm-6 hisG4 rfbD1 R(zgb210::Tn10) TetS endA1 rspL136 (StrR ) dam13::Tn9 (CamR ) xylA-5 mtl-1 thi-1 mcrB1 hsdR2. .. Bacillus anthracis (Sterne 34 F2) was obtained from Colorado Serum Co. (Denver, CO).

    Recombinant:

    Article Title: Target-dependent nickase activities of CRISPR-Cas nucleases Cpf1 and Cas9
    Article Snippet: .. Recombinant proteins were expressed in modified E. coli NiCo21 (DE3) cells (NEB C2925H) harboring the Cpf1 expression plasmid by growing in LB at 23°C for 16 hr in presence of IPTG at 0.4mM. .. As/Fn/Lb-Cpf1 was purified using HiTrap DEAE FF (GE Healthcare), HisTrap HP (Ni-NTA) (GE Healthcare) and HiTrapSP HP (GE Healthcare) columns.

    Homologous Recombination:

    Article Title: Chlamydia trachomatis transformation and allelic exchange mutagenesis
    Article Snippet: .. pSUmC plasmid DNA pUC18A plasmid DNA pUC19G plasmid DNA Primer pair to amplify the gene targeted for deletion surrounded by approximately 3-kilobase (kb) flanking sequences from chlamydial genomic DNA: X@pUC F and R Primer pair surrounding, but directed away from, the target gene: X-away F and R Primer pair for amplifying the bla-gfp cassette from pUC19G: bla-gfp F: GGAAATGTGCGCGGAACCC bla-gfp R: TTACTTGTATAGTTCATCCATGCCATGTG Primer pair for amplifying the homologous recombination sequence for insertion into pSUmC: HomRR@pSUmC F: CTGCAGGTACCGGTCGACCATTC GGTCTGACGCTCAGTGGAACG HomRR@pSUmC R: GATCTTTCTACGGGGTCTGACGCTC CTGGCGTTACCCAACTTAATCGCC Q5® High-Fidelity DNA Polymerase (M0491S, NEB) DpnI restriction enzyme (R0176S, NEB) Cutsmart buffer (B7204S, NEB) T4 Polynucleotide Kinase (M0201S, NEB) Quick Ligation™ Kit (M2200S, NEB) NEB® 10-beta Competent E. coli (C3019H, NEB) dam− /dcm− Competent E. coli (C2925I, NEB) Zyppy™ Plasmid Miniprep Kit (D4036, Zymo Research) Zymoclean™ Gel DNA Recovery Kit (D4001, Zymo Research) QIAfilter Plasmid Maxi Kit (12262, Qiagen) Spectinomycin dihydrochloride pentahydrate (J61820, Alfa Aesar) Carbenicillin disodium salt (J61949, Alfa Aesar) LB Media (J106, VWR) Agar Powder ( , Alfa Aesar) Agarose LE (BMK-A1705, KSE Scientific) Phenol–Chloroform (6805, EMD Millipore) 3M sodium acetate 100% ethanol Design primers for amplifying gene × flanked upstream and downstream (±) by 3 kb arms for insertion into pUC18A. .. Sequences should be designed with melting temperatures of approximately 60°C which hairpin at temperatures no higher than 40°C, as determined by OligoAnalyzer 3.1 (Integrated DNA Technologies).

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    New England Biolabs dam dcm competent e coli
    Schematic of overall cloning strategy (Basic Protocol 1) used for constructing suicide plasmids. 1. The gene of interest ( ct00X ) is amplified from C. trachomatis L2 genomic DNA with 3 kb of up (us)- and down (ds)-stream homology arm (HR) DNA (steps 1–3). 2. The PCR product is used in an insertion/deletion PCR reaction using pUC18A as template such that the chlamydial DNA sequence replaces plasmid-encoded bla . (steps 4–7). 3. The target gene ( ct00X ) is deleted using Away primers, annealing immediately outside the ct00X coding sequence, in a divergent PCR reaction to amplify the remaining pUC18@ct00X construct. The divergent PCR product is then ligated to the bla and gfp cassette amplified from pUC19G to generate a pUC18 plasmid where ct00X has been replaced by bla-gfp (steps 8–13). 4. The chlamydial DNA required for homologous recombination plus the bla-gfp selection marker is PCR amplified using engineered primers (Step 14). 5. An insertion PCR is used to replace the bla gene in pSuMc with the homologous-recombination construct (steps 15–18). 6. The completed suicide plasmid is propagated in dam − <t>/dcm</t> − E. coli prior to the transformation of chlamydiae outlined in protocol 2 (step 19).
    Dam Dcm Competent E Coli, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dam dcm competent e coli/product/New England Biolabs
    Average 95 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    dam dcm competent e coli - by Bioz Stars, 2020-05
    95/100 stars
      Buy from Supplier

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    Schematic of overall cloning strategy (Basic Protocol 1) used for constructing suicide plasmids. 1. The gene of interest ( ct00X ) is amplified from C. trachomatis L2 genomic DNA with 3 kb of up (us)- and down (ds)-stream homology arm (HR) DNA (steps 1–3). 2. The PCR product is used in an insertion/deletion PCR reaction using pUC18A as template such that the chlamydial DNA sequence replaces plasmid-encoded bla . (steps 4–7). 3. The target gene ( ct00X ) is deleted using Away primers, annealing immediately outside the ct00X coding sequence, in a divergent PCR reaction to amplify the remaining pUC18@ct00X construct. The divergent PCR product is then ligated to the bla and gfp cassette amplified from pUC19G to generate a pUC18 plasmid where ct00X has been replaced by bla-gfp (steps 8–13). 4. The chlamydial DNA required for homologous recombination plus the bla-gfp selection marker is PCR amplified using engineered primers (Step 14). 5. An insertion PCR is used to replace the bla gene in pSuMc with the homologous-recombination construct (steps 15–18). 6. The completed suicide plasmid is propagated in dam − /dcm − E. coli prior to the transformation of chlamydiae outlined in protocol 2 (step 19).

    Journal: Current protocols in microbiology

    Article Title: Chlamydia trachomatis transformation and allelic exchange mutagenesis

    doi: 10.1002/cpmc.31

    Figure Lengend Snippet: Schematic of overall cloning strategy (Basic Protocol 1) used for constructing suicide plasmids. 1. The gene of interest ( ct00X ) is amplified from C. trachomatis L2 genomic DNA with 3 kb of up (us)- and down (ds)-stream homology arm (HR) DNA (steps 1–3). 2. The PCR product is used in an insertion/deletion PCR reaction using pUC18A as template such that the chlamydial DNA sequence replaces plasmid-encoded bla . (steps 4–7). 3. The target gene ( ct00X ) is deleted using Away primers, annealing immediately outside the ct00X coding sequence, in a divergent PCR reaction to amplify the remaining pUC18@ct00X construct. The divergent PCR product is then ligated to the bla and gfp cassette amplified from pUC19G to generate a pUC18 plasmid where ct00X has been replaced by bla-gfp (steps 8–13). 4. The chlamydial DNA required for homologous recombination plus the bla-gfp selection marker is PCR amplified using engineered primers (Step 14). 5. An insertion PCR is used to replace the bla gene in pSuMc with the homologous-recombination construct (steps 15–18). 6. The completed suicide plasmid is propagated in dam − /dcm − E. coli prior to the transformation of chlamydiae outlined in protocol 2 (step 19).

    Article Snippet: pSUmC plasmid DNA pUC18A plasmid DNA pUC19G plasmid DNA Primer pair to amplify the gene targeted for deletion surrounded by approximately 3-kilobase (kb) flanking sequences from chlamydial genomic DNA: X@pUC F and R Primer pair surrounding, but directed away from, the target gene: X-away F and R Primer pair for amplifying the bla-gfp cassette from pUC19G: bla-gfp F: GGAAATGTGCGCGGAACCC bla-gfp R: TTACTTGTATAGTTCATCCATGCCATGTG Primer pair for amplifying the homologous recombination sequence for insertion into pSUmC: HomRR@pSUmC F: CTGCAGGTACCGGTCGACCATTC GGTCTGACGCTCAGTGGAACG HomRR@pSUmC R: GATCTTTCTACGGGGTCTGACGCTC CTGGCGTTACCCAACTTAATCGCC Q5® High-Fidelity DNA Polymerase (M0491S, NEB) DpnI restriction enzyme (R0176S, NEB) Cutsmart buffer (B7204S, NEB) T4 Polynucleotide Kinase (M0201S, NEB) Quick Ligation™ Kit (M2200S, NEB) NEB® 10-beta Competent E. coli (C3019H, NEB) dam− /dcm− Competent E. coli (C2925I, NEB) Zyppy™ Plasmid Miniprep Kit (D4036, Zymo Research) Zymoclean™ Gel DNA Recovery Kit (D4001, Zymo Research) QIAfilter Plasmid Maxi Kit (12262, Qiagen) Spectinomycin dihydrochloride pentahydrate (J61820, Alfa Aesar) Carbenicillin disodium salt (J61949, Alfa Aesar) LB Media (J106, VWR) Agar Powder ( , Alfa Aesar) Agarose LE (BMK-A1705, KSE Scientific) Phenol–Chloroform (6805, EMD Millipore) 3M sodium acetate 100% ethanol Design primers for amplifying gene × flanked upstream and downstream (±) by 3 kb arms for insertion into pUC18A.

    Techniques: Clone Assay, Amplification, Polymerase Chain Reaction, Sequencing, Plasmid Preparation, Construct, Homologous Recombination, Selection, Marker, Transformation Assay