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RSF1010-based shuttle vectors for cloning and expression in Pasteurella multocida

M D Lee1, A D Henk

  • 1College of Veterinary Medicine, University of Georgia, Athens 30601, USA. lee.m@calc.vet.uga.edu

Veterinary Microbiology
|March 1, 1997
PubMed
Summary

Broad host-range vectors pJRD215 and pMMB67EH show promise for Pasteurella multocida cloning. Conjugation proved efficient for transfer, enabling gene expression and functional protein production in P. multocida.

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Area of Science:

  • Molecular Biology
  • Microbiology
  • Genetics

Background:

  • Pasteurella multocida is a significant animal pathogen.
  • Efficient genetic manipulation tools are crucial for studying P. multocida.
  • Broad host-range vectors offer versatile cloning capabilities.

Purpose of the Study:

  • To assess the stability and cloning efficiency of pJRD215 and pMMB67EH vectors in P. multocida.
  • To evaluate different DNA transfer methods into P. multocida.
  • To determine the functional expression of cloned genes in P. multocida.

Main Methods:

  • Electroporation and conjugation were used for DNA transfer.
  • Vector stability was assessed in P. multocida.
  • Gene expression (LacZ) was confirmed via functional assays (ONPG test).

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Main Results:

  • Electroporation yielded unreliable and inefficient transformation of P. multocida.
  • Conjugation from E. coli enabled high-efficiency transfer of both vectors.
  • Antibiotic resistance markers and LacZ gene expression were confirmed in P. multocida transconjugants.

Conclusions:

  • pJRD215 and pMMB67EH are effective shuttle vectors for cloning in P. multocida.
  • Conjugation is a superior method for introducing these vectors into P. multocida.
  • These vectors facilitate genetic studies and manipulation of P. multocida.