Home LiteratureArticle Details
PMID: 3018431 Published · ppublish English Journal Article

Construction and characterisation of a series of multi-copy promoter-probe plasmid vectors for Streptomyces using the aminoglycoside phosphotransferase gene from Tn5 as indicator.

Molecular & general genetics : MGG ·Vol. 203 ·No. 3 ·1986-06-00 ·Pages 468-78

Ward JM, Janssen GR, Kieser T, Bibb MJ, Buttner MJ, Bibb MJ

Abstract

Several versatile, multi-copy, promoter-probe plasmid vectors have been constructed that replicate in a wide range of Streptomyces species. Transcriptional activity is detected by the expression of a promoter-less aminoglycoside phosphotransferase gene (neo) derived from the transposon Tn5; expression of this gene confers kanamycin and neomycin resistance on Streptomyces lividans. An efficient transcriptional terminator from E. coli phage fd has been inserted upstream of the neo coding region to prevent significant transcriptional read-through from vector promoters. A translational stop codon situated downstream from the site(s) used for cloning and preceding and in frame with the ATG start codon of the neo gene ensures the detection of transcriptional, rather than translational, fusions. Relative promoter strengths can be determined by gradient plate assays of kanamycin resistance, by measuring the amount of aminoglycoside phosphotransferase produced or by estimating neo mRNA synthesised. The high copy number of the vectors facilitates the rapid isolation and characterisation of promoter-active fragments and convenient restriction sites are available for DNA sequencing and S1 mapping of cloned inserts. Some derivatives contain a polylinker that facilitates the insertion, excision and analysis of cloned fragments and which enhances the use of these plasmids as general cloning vectors.

MeSH Terms
Cloning, Molecular DNA Restriction Enzymes DNA Transposable Elements Genes Genes, Bacterial Genetic Vectors Kanamycin Kinase Nucleic Acid Hybridization Phosphotransferases/genetics Plasmids Promoter Regions, Genetic Streptomyces/enzymology,genetics
Chemicals
DNA Transposable Elements Phosphotransferases Kanamycin Kinase DNA Restriction Enzymes
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Ward J M
Janssen G R
Kieser T
Bibb M J
Buttner M J
Bibb M J
References (45)
45 references, click to expand
  1. Construction of biologically functional bacterial plasmids in vitro.
    Proc Natl Acad Sci U S A. 1973 Nov;70(11):3240-4 PMID: 4594039
  2. Regulatory sequences involved in the promotion and termination of RNA transcription.
    Annu Rev Genet. 1979;13:319-53 PMID: 94251
  3. Phage-mediated cloning of bldA, a region involved in Streptomyces coelicolor morphological development, and its analysis by genetic complementation.
    J Bacteriol. 1985 Sep;163(3):965-72 PMID: 2993254
  4. pIJ101, a multi-copy broad host-range Streptomyces plasmid: functional analysis and development of DNA cloning vectors.
    Mol Gen Genet. 1982;185(2):223-8 PMID: 6283316
  5. Rapid and efficient cosmid cloning.
    Nucleic Acids Res. 1981 Jul 10;9(13):2989-98 PMID: 6269067
  6. Construction and application of a promoter-probe plasmid that allows chromogenic identification in Streptomyces lividans.
    J Bacteriol. 1985 Apr;162(1):406-12 PMID: 2984181
  7. New versatile plasmid vectors for expression of hybrid proteins coded by a cloned gene fused to lacZ gene sequences encoding an enzymatically active carboxy-terminal portion of beta-galactosidase.
    Gene. 1983 Nov;25(1):71-82 PMID: 6319233
  8. Streptomyces lividans RNA polymerase recognizes and uses Escherichia coli transcriptional signals.
    Gene. 1984 Apr;28(1):83-91 PMID: 6203812
  9. The glycerol utilization operon of Streptomyces coelicolor: genetic mapping of gyl mutations and the analysis of cloned gylDNA.
    Mol Gen Genet. 1984;193(1):119-28 PMID: 6318046
  10. Analysis of gene control signals by DNA fusion and cloning in Escherichia coli.
    J Mol Biol. 1980 Apr;138(2):179-207 PMID: 6997493
  11. The nucleotide sequence of the tyrosinase gene from Streptomyces antibioticus and characterization of the gene product.
    Gene. 1985;37(1-3):101-10 PMID: 3932128
  12. The Streptomyces plasmid SCP2*: its functional analysis and development into useful cloning vectors.
    Gene. 1985;35(3):223-35 PMID: 2995202
  13. Nonchromosomal antibiotic resistance in bacteria: genetic transformation of Escherichia coli by R-factor DNA.
    Proc Natl Acad Sci U S A. 1972 Aug;69(8):2110-4 PMID: 4559594
  14. RNA polymerase heterogeneity in Streptomyces coelicolor.
    Nature. 1985 Jan 3-9;313(5997):22-7 PMID: 2578216
  15. RNA polymerase-DNA interactions in Streptomyces. In vitro studies of a S. lividans plasmid promoter with S. coelicolor RNA polymerase.
    J Mol Biol. 1985 Sep 5;185(1):177-88 PMID: 4046037
  16. Molecular cloning of the whole biosynthetic pathway of a Streptomyces antibiotic and its expression in a heterologous host.
    Nature. 1984 May 31-Jun 6;309(5967):462-4 PMID: 6328317
  17. Isolation of deoxyribonucleic acid and ribosomal ribonucleic acid from bacteria.
    Biochem J. 1967 Jul;104(1):258-62 PMID: 4962318
  18. Cloning and analysis of the promoter region of the erythromycin resistance gene (ermE) of Streptomyces erythraeus.
    Gene. 1985;38(1-3):215-26 PMID: 2998943
  19. Microbial Selection.
    Science. 1952 Jul 18;116(3003):45-51 PMID: 17813331
  20. Cloning and expression in streptomyces lividans of antibiotic resistance genes derived from Escherichia coli.
    J Bacteriol. 1981 Apr;146(1):360-8 PMID: 7012122
  21. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers.
    Gene. 1982 Oct;19(3):259-68 PMID: 6295879
  22. Factors affecting the isolation of CCC DNA from Streptomyces lividans and Escherichia coli.
    Plasmid. 1984 Jul;12(1):19-36 PMID: 6387733
  23. The use of thin acrylamide gels for DNA sequencing.
    FEBS Lett. 1978 Mar 1;87(1):107-10 PMID: 631324
  24. Cloning and analysis of strong promoters is made possible by the downstream placement of a RNA termination signal.
    Proc Natl Acad Sci U S A. 1981 Aug;78(8):4936-40 PMID: 6946440
  25. Cloning and characterization of EcoRI and HindIII restriction endonuclease-generated fragments of antibiotic resistance plasmids R6-5 and R6.
    Mol Gen Genet. 1978 Jun 14;162(2):121-37 PMID: 672900
  26. Plasmids, recombination and chromosome mapping in Streptomyces lividans 66.
    J Gen Microbiol. 1983 Jul;129(7):2257-69 PMID: 6631413
  27. Genetic analysis and genome structure in Streptomyces coelicolor.
    Bacteriol Rev. 1967 Dec;31(4):373-403 PMID: 4866846
  28. pUR222, a vector for cloning and rapid chemical sequencing of DNA.
    Nucleic Acids Res. 1981 Aug 25;9(16):4087-98 PMID: 6272193
  29. DNAGEL: a computer program for determining DNA fragment sizes using a small computer equipped with a graphics tablet.
    Nucleic Acids Res. 1984 Jan 11;12(1 Pt 2):679-88 PMID: 6320102
  30. A rapid alkaline extraction procedure for screening recombinant plasmid DNA.
    Nucleic Acids Res. 1979 Nov 24;7(6):1513-23 PMID: 388356
  31. Transduction of linked genetic characters of the host by bacteriophage P1.
    Virology. 1955 Jul;1(2):190-206 PMID: 13267987
  32. Transcription maps of polyoma virus-specific RNA: analysis by two-dimensional nuclease S1 gel mapping.
    Methods Enzymol. 1980;65(1):718-49 PMID: 6154876
  33. Advances in Streptomyces coelicolor genetics.
    Bacteriol Rev. 1973 Sep;37(3):371-405 PMID: 4585795
  34. Nucleotide sequences encoding and promoting expression of three antibiotic resistance genes indigenous to Streptomyces.
    Mol Gen Genet. 1985;199(1):26-36 PMID: 2987648
  35. Physical analysis of antibiotic-resistance genes from Streptomyces and their use in vector construction.
    Gene. 1982 Nov;20(1):51-62 PMID: 6298066
  36. Genetic recombination through protoplast fusion in Streptomyces.
    Nature. 1977 Jul 14;268(5616):171-4 PMID: 593313
  37. Cloning and expression of the tyrosinase gene from Streptomyces antibioticus in Streptomyces lividans.
    J Gen Microbiol. 1983 Sep;129(9):2703-14 PMID: 6313861
  38. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.
    Anal Biochem. 1976 May 7;72:248-54 PMID: 942051
  39. Gene cloning in Streptomyces.
    Curr Top Microbiol Immunol. 1982;96:69-95 PMID: 6276096
  40. Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis.
    Gene. 1983 Dec;26(1):101-6 PMID: 6323249
  41. Sequencing end-labeled DNA with base-specific chemical cleavages.
    Methods Enzymol. 1980;65(1):499-560 PMID: 6246368
  42. Nucleotide sequence and exact localization of the neomycin phosphotransferase gene from transposon Tn5.
    Gene. 1982 Oct;19(3):327-36 PMID: 6295884
  43. Gene expression in Streptomyces: construction and application of promoter-probe plasmid vectors in Streptomyces lividans.
    Mol Gen Genet. 1982;187(2):265-77 PMID: 6294463
  44. Cloning in single-stranded bacteriophage as an aid to rapid DNA sequencing.
    J Mol Biol. 1980 Oct 25;143(2):161-78 PMID: 6260957
  45. Resistance, regulatory and production genes for the antibiotic methylenomycin are clustered.
    EMBO J. 1985 Jul;4(7):1893-7 PMID: 2992952
Article Info
Journal
Molecular & general genetics : MGG
Abbr.
Mol Gen Genet
ISSN
0026-8925
DOI
10.1007/BF00422072
Published
1986-06-00
Pages
468-78
Language
English
Region
Germany
NLM ID
0125036
Subset
IM
External Links
PubMed source DOI: 10.1007/BF00422072
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2