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Related Experiment Videos

Plasmid pIJ699, a multi-copy positive-selection vector for Streptomyces.

T Kieser1, R E Melton

  • 1John Innes Institute, Norwich, U.K.

Gene
|May 15, 1988
PubMed
Summary
This summary is machine-generated.

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A new plasmid vector, pIJ699, enables positive selection for cloned DNA in bacteria. This novel system enhances plasmid stability and simplifies gene regulation studies in Streptomyces.

Area of Science:

  • Molecular Biology
  • Microbial Genetics
  • Biotechnology

Background:

  • Plasmid vectors are essential tools in molecular biology for DNA cloning and gene expression.
  • Developing vectors with robust selection mechanisms is crucial for efficient genetic manipulation.
  • Streptomyces species are important for antibiotic production, necessitating effective genetic tools.

Purpose of the Study:

  • To construct a novel plasmid vector, pIJ699, for positive selection of cloned DNA.
  • To leverage the replication functions of the broad-host-range plasmid pIJ101 for Streptomyces applications.
  • To enhance the stability of hybrid plasmids and facilitate the study of gene regulation.

Main Methods:

  • Construction of pIJ699 using replication functions from pIJ101.

Related Experiment Videos

  • Utilizing the principle that plasmids with uninterrupted inverted repeats are non-viable for selection.
  • Digestion with BglII, separation of vector and spacer fragments, and ligation with target DNA.
  • Selection of thiostrepton-resistant transformants in Streptomyces lividans.

    • Main Results:

    • pIJ699 successfully provides positive selection for cloned DNA inserts.
    • The vector design, incorporating inverted repeats and a transcription terminator, enhances plasmid stability.
    • Reduced transcriptional read-through improves the study of cloned gene regulation.
    • Successful cloning and selection demonstrated in Streptomyces lividans.

    Conclusions:

    • The pIJ699 plasmid vector offers an effective positive selection system for DNA cloning in Streptomyces.
    • This vector enhances hybrid plasmid stability and facilitates genetic studies.
    • pIJ699 represents a valuable tool for molecular biology research involving Streptomyces.