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puc19  (Valiant Co Ltd)
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Replacement in B. subtilis ATCC 6633 of the original Pmyc promoter by the PrepU-neo cassette using homologous recombination between genomic DNA of the strain and hybrid plasmid pBG106. (A) Recognition of homologous regions located (i) after the termination region of the pbp gene (coding for a penicillin-binding protein) located upstream of the mycosubtilin operon (for convenience, the cassette generated by PCR in this region was designated ɛ“pbp”) and (ii) immediately downstream of the Pmyc promoter (cassette ɛfenF). Four genes, fenF, mycA, mycB and mycC, constitute the mycosubtilin operon and code for a malonyl coenzyme A transacylase and three peptide synthetases, respectively. yngL, gene coding for an unknown function; PrepU, promoter of the replication gene of pUB110; neo, gene conferring resistance to neomycin/kanamycin from pUB110 (15). An asterisk indicates the site newly created after ligation between the BspEI- and XmaI-compatible cohesive ends. (B) Construct obtained for the genomic DNA of the strain following homologous recombination (generated by the inability of <t>pUC19</t> to replicate in Bacillus spp., together with the selective pressure for resistance to neomycin). The mycosubtilin operon came under control of the PrepU constitutive promoter.
Puc19, supplied by Valiant Co Ltd, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/04821227/pUC19+plasmid/pmc01183317-167-4-31
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Replacement in B. subtilis ATCC 6633 of the original Pmyc promoter by the PrepU-neo cassette using homologous recombination between genomic DNA of the strain and hybrid plasmid pBG106. (A) Recognition of homologous regions located (i) after the termination region of the pbp gene (coding for a penicillin-binding protein) located upstream of the mycosubtilin operon (for convenience, the cassette generated by PCR in this region was designated ɛ“pbp”) and (ii) immediately downstream of the Pmyc promoter (cassette ɛfenF). Four genes, fenF, mycA, mycB and mycC, constitute the mycosubtilin operon and code for a malonyl coenzyme A transacylase and three peptide synthetases, respectively. yngL, gene coding for an unknown function; PrepU, promoter of the replication gene of pUB110; neo, gene conferring resistance to neomycin/kanamycin from pUB110 (15). An asterisk indicates the site newly created after ligation between the BspEI- and XmaI-compatible cohesive ends. (B) Construct obtained for the genomic DNA of the strain following homologous recombination (generated by the inability of pUC19 to replicate in Bacillus spp., together with the selective pressure for resistance to neomycin). The mycosubtilin operon came under control of the PrepU constitutive promoter.

Journal:

Article Title: Mycosubtilin Overproduction by Bacillus subtilis BBG100 Enhances the Organism's Antagonistic and Biocontrol Activities

doi: 10.1128/AEM.71.8.4577-4584.2005

Figure Lengend Snippet: Replacement in B. subtilis ATCC 6633 of the original Pmyc promoter by the PrepU-neo cassette using homologous recombination between genomic DNA of the strain and hybrid plasmid pBG106. (A) Recognition of homologous regions located (i) after the termination region of the pbp gene (coding for a penicillin-binding protein) located upstream of the mycosubtilin operon (for convenience, the cassette generated by PCR in this region was designated ɛ“pbp”) and (ii) immediately downstream of the Pmyc promoter (cassette ɛfenF). Four genes, fenF, mycA, mycB and mycC, constitute the mycosubtilin operon and code for a malonyl coenzyme A transacylase and three peptide synthetases, respectively. yngL, gene coding for an unknown function; PrepU, promoter of the replication gene of pUB110; neo, gene conferring resistance to neomycin/kanamycin from pUB110 (15). An asterisk indicates the site newly created after ligation between the BspEI- and XmaI-compatible cohesive ends. (B) Construct obtained for the genomic DNA of the strain following homologous recombination (generated by the inability of pUC19 to replicate in Bacillus spp., together with the selective pressure for resistance to neomycin). The mycosubtilin operon came under control of the PrepU constitutive promoter.

Article Snippet: For construction of the pUC19-derived plasmid dedicated to promoter exchange by homologous recombination in B. subtilis , the pbp and fenF fragments were generated by PCR using Taq polymerase “Arrow” from Qbiogene (Montreal, Canada).

Techniques: Homologous Recombination, Plasmid Preparation, Binding Assay, Generated, Ligation, Construct

Strains and plasmids

Journal:

Article Title: Mycosubtilin Overproduction by Bacillus subtilis BBG100 Enhances the Organism's Antagonistic and Biocontrol Activities

doi: 10.1128/AEM.71.8.4577-4584.2005

Figure Lengend Snippet: Strains and plasmids

Article Snippet: For construction of the pUC19-derived plasmid dedicated to promoter exchange by homologous recombination in B. subtilis , the pbp and fenF fragments were generated by PCR using Taq polymerase “Arrow” from Qbiogene (Montreal, Canada).

Techniques: Plasmid Preparation, Clone Assay