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Construction of versatile low-copy-number vectors for cloning, sequencing and gene expression in Escherichia coli.
Gene 1991; 100:195-9GENE

Abstract

Using the polymerase chain reaction and standard recombinant DNA techniques, a series of new multipurpose low-copy-number (lcn) vectors, pWSK29, pWKS30, pWKS129 and pWKS130, have been constructed. Plasmids pWSK29 and pWKS30 carry the ampicillin-resistance marker (ApR), 20 unique cloning sites flanked by T7 and T3 RNA polymerase promoters, the lacZ alpha gene and the bacteriophage f1 origin of replication (ori) for production of single-stranded (ss) DNA in the presence of a helper phage. Plasmids pWSK129 and pWKS130 carry the kanamycin-resistance marker (KmR) and have 16 unique cloning sites flanked by T7 and T3 RNA polymerase promoters positioned within the lacZ alpha gene. Plasmids pWSK129 and pWKS130 also contain the f1 ori for the generation of ss DNA in the presence of a helper phage. All of the plasmids have an lcn of six to eight per cell. Each vector can be used for: (i) complementation analysis, (ii) generating unidirectional deletions with exonuclease III/S1 nuclease, (iii) DNA sequencing, (iv) high-level gene expression using T7 RNA polymerase, and (v) run-off transcription. They are very useful for analyzing genes encoding proteins which are toxic in Escherichia coli in high copy number.

Authors+Show Affiliations

Department of Genetics, University of Georgia, Athens 30602.No affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

2055470

Citation

Wang, R F., and S R. Kushner. "Construction of Versatile Low-copy-number Vectors for Cloning, Sequencing and Gene Expression in Escherichia Coli." Gene, vol. 100, 1991, pp. 195-9.
Wang RF, Kushner SR. Construction of versatile low-copy-number vectors for cloning, sequencing and gene expression in Escherichia coli. Gene. 1991;100:195-9.
Wang, R. F., & Kushner, S. R. (1991). Construction of versatile low-copy-number vectors for cloning, sequencing and gene expression in Escherichia coli. Gene, 100, pp. 195-9.
Wang RF, Kushner SR. Construction of Versatile Low-copy-number Vectors for Cloning, Sequencing and Gene Expression in Escherichia Coli. Gene. 1991;100:195-9. PubMed PMID: 2055470.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Construction of versatile low-copy-number vectors for cloning, sequencing and gene expression in Escherichia coli. AU - Wang,R F, AU - Kushner,S R, PY - 1991/4/1/pubmed PY - 1991/4/1/medline PY - 1991/4/1/entrez SP - 195 EP - 9 JF - Gene JO - Gene VL - 100 N2 - Using the polymerase chain reaction and standard recombinant DNA techniques, a series of new multipurpose low-copy-number (lcn) vectors, pWSK29, pWKS30, pWKS129 and pWKS130, have been constructed. Plasmids pWSK29 and pWKS30 carry the ampicillin-resistance marker (ApR), 20 unique cloning sites flanked by T7 and T3 RNA polymerase promoters, the lacZ alpha gene and the bacteriophage f1 origin of replication (ori) for production of single-stranded (ss) DNA in the presence of a helper phage. Plasmids pWSK129 and pWKS130 carry the kanamycin-resistance marker (KmR) and have 16 unique cloning sites flanked by T7 and T3 RNA polymerase promoters positioned within the lacZ alpha gene. Plasmids pWSK129 and pWKS130 also contain the f1 ori for the generation of ss DNA in the presence of a helper phage. All of the plasmids have an lcn of six to eight per cell. Each vector can be used for: (i) complementation analysis, (ii) generating unidirectional deletions with exonuclease III/S1 nuclease, (iii) DNA sequencing, (iv) high-level gene expression using T7 RNA polymerase, and (v) run-off transcription. They are very useful for analyzing genes encoding proteins which are toxic in Escherichia coli in high copy number. SN - 0378-1119 UR - https://www.unboundmedicine.com/medline/citation/2055470/Construction_of_versatile_low_copy_number_vectors_for_cloning_sequencing_and_gene_expression_in_Escherichia_coli_ L2 - https://linkinghub.elsevier.com/retrieve/pii/0378-1119(91)90366-J DB - PRIME DP - Unbound Medicine ER -