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Related Experiment Videos

Stability function in the Bacillus subtilis plasmid pTA 1060.

S Bron1, P Bosma, M van Belkum

  • 1Department of Genetics, Center of Biological Sciences, Haren, The Netherlands.

Plasmid
|July 1, 1987
PubMed
Summary
This summary is machine-generated.

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Researchers developed a stable Bacillus subtilis plasmid, pBB2, for genetic engineering. Deleting specific DNA fragments from pBB2 resulted in instability, indicating a crucial stability element within those fragments.

Area of Science:

  • Molecular Biology
  • Microbial Genetics

Background:

  • The Bacillus subtilis cryptic plasmid pTA 1060 was genetically modified.
  • Chloramphenicol resistance (CmR) and kanamycin resistance (KmR) markers were introduced.

Purpose of the Study:

  • To construct and characterize a stable plasmid vector for Bacillus subtilis.
  • To identify genetic elements responsible for plasmid stability.

Main Methods:

  • Construction of plasmid pBB2 by integrating CmR and KmR markers into pTA 1060.
  • Deletion mutagenesis to create pBB3 and assess stability.
  • Analysis of plasmid loss rates in nonselective media.
  • Evaluation of copy number effects on stability.

Main Results:

Related Experiment Videos

  • Plasmid pBB2 exhibited high segregation stability (loss rate ≤ 0.02% per cell generation).
  • Deletion of specific ClaI fragments (1.45 and 0.20 kb) to create pBB3 resulted in significant instability (loss rate ≥ 0.5% per cell generation).
  • Increased copy number in pBB3 cop mutant partially restored stability, suggesting a link between copy number and stability.

    • Conclusions:

    • A genetic element(s) essential for plasmid stability is located on the deleted ClaI fragments.
    • The constructed pBB2 plasmid demonstrates potential utility as a cloning vector for large DNA fragments in Bacillus subtilis.