PSET152: Difference between revisions
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{{DISPLAYTITLE: pSET152}} | {{DISPLAYTITLE: pSET152}} | ||
==Features== | |||
• Backbone: Derived from a standard E. coli plasmid vector (pBR322/pUC lineage) containing an E. coli replication origin (ori) and an origin of transfer (oriT) derived from broad-host-range plasmid RK2 to allow conjugal transfer. | |||
• Integrative Machinery: Incorporates the attachment site (attP) and the integrase gene (int) from the temperate actinophage φC31. Integrates into the φC31 att site on the Streptomyces genome. | |||
• Selection Marker: Carries the aac(3)IV gene, which confers resistance to apramycin in both E. coli and Streptomyces. | |||
==History== | ==History== | ||
The pSET152 plasmid was constructed and published by Bierman et al. in 1992 as an E. coli–Streptomyces shuttle and integrative vector | |||
==Map== | ==Map== | ||
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==References== | ==References== | ||
M. Bierman, R. Logan, K. O'Brien, E.T. Seno, R. Nagaraja Rao, B.E. Schoner (1992). Plasmid cloning vectors for the conjugal transfer of DNA from Escherichia coli to Streptomyces spp. Gene 116:43-49. | |||
Abstract | |||
We have constructed cloning vectors for the conjugal transfer of DNA from Escherichia coli to Streptomyces spp. All vectors contain the 760-bp oriT fragment from the IncP plasmid, RK2. Transfer functions need to be supplied in trans by the E. coli donor strain. We have incorporated into these vectors selectable antibiotic-resistance markers (AmR, ThR, SpR) that function in Streptomyces spp. and other features that should allow for: (i) integration via homologous recombination between cloned DNA and the Streptomyces spp. chromosome, (ii) autonomous replication, or (iii) site-specific integration at the bacteriophage φC31 attachment site. Shuttle cosmids for constructing genomic libraries and bacteriophage P1 cloning vector capable of accepting approx. 100-kb fragments are also described. A simple mating procedure has been developed for the conjugal transfer of these vectors from E. coli to Streptomyces spp. that involves plating of the donor strain and either germinated spores or mycelial fragments of the recipient strain. We have shown that several of these vectors can be introduced into Streptomyces fradiae, a strain that is notoriously difficult to transform by PEG-mediated protoplast transformation. | |||
https://www.sciencedirect.com/science/article/abs/pii/0378111992906272 | |||
Latest revision as of 09:28, 10 October 2026
Features
• Backbone: Derived from a standard E. coli plasmid vector (pBR322/pUC lineage) containing an E. coli replication origin (ori) and an origin of transfer (oriT) derived from broad-host-range plasmid RK2 to allow conjugal transfer.
• Integrative Machinery: Incorporates the attachment site (attP) and the integrase gene (int) from the temperate actinophage φC31. Integrates into the φC31 att site on the Streptomyces genome.
• Selection Marker: Carries the aac(3)IV gene, which confers resistance to apramycin in both E. coli and Streptomyces.
History
The pSET152 plasmid was constructed and published by Bierman et al. in 1992 as an E. coli–Streptomyces shuttle and integrative vector
Map
Sequence links
References
M. Bierman, R. Logan, K. O'Brien, E.T. Seno, R. Nagaraja Rao, B.E. Schoner (1992). Plasmid cloning vectors for the conjugal transfer of DNA from Escherichia coli to Streptomyces spp. Gene 116:43-49.
Abstract We have constructed cloning vectors for the conjugal transfer of DNA from Escherichia coli to Streptomyces spp. All vectors contain the 760-bp oriT fragment from the IncP plasmid, RK2. Transfer functions need to be supplied in trans by the E. coli donor strain. We have incorporated into these vectors selectable antibiotic-resistance markers (AmR, ThR, SpR) that function in Streptomyces spp. and other features that should allow for: (i) integration via homologous recombination between cloned DNA and the Streptomyces spp. chromosome, (ii) autonomous replication, or (iii) site-specific integration at the bacteriophage φC31 attachment site. Shuttle cosmids for constructing genomic libraries and bacteriophage P1 cloning vector capable of accepting approx. 100-kb fragments are also described. A simple mating procedure has been developed for the conjugal transfer of these vectors from E. coli to Streptomyces spp. that involves plating of the donor strain and either germinated spores or mycelial fragments of the recipient strain. We have shown that several of these vectors can be introduced into Streptomyces fradiae, a strain that is notoriously difficult to transform by PEG-mediated protoplast transformation.
https://www.sciencedirect.com/science/article/abs/pii/0378111992906272
