Sequence analysis of a cryptic plasmid pCJ419 from Campylobacter jejuni and construction of an Escherichia coli-Campylobacter shuttle vector

Plasmid. 2003 Sep;50(2):152-60. doi: 10.1016/s0147-619x(03)00060-x.

Abstract

A small cryptic plasmid, pCJ419, was identified in a human clinical isolate of Campylobacter jejuni, cloned and sequenced. pCJ419 is a circular molecule of 4013 bp with a G+C content of 27.1%. The products of four open reading frames (ORFs) share significant sequence similarity with putative proteins from known C. jejuni and Campylobacter coli plasmids. ORF-1 encodes a putative mobilisation protein (Mob). ORF-2 and ORF-3 encode proteins that have high identity to putative RepA and RepB proteins, respectively, of known C. jejuni and C. coli plasmids. ORF-4 encodes a protein that has high identity to a hypothetical protein of unknown function, Cjp32, previously described in a pVir plasmid of C. jejuni. Tandem repeating 22-bp sequences typical of a plasmid replication origin (ori) were identified upstream of the DNA sequences encoding putative replication initiation proteins. An Escherichia coli-Campylobacter shuttle cloning vector, pGU0202, was constructed using plasmid pMW2 that harbours a Campylobacter-derived kanamycin resistance gene [aph(3')-III]. The sequences encoding pCJ419 mob, RepA and RepB proteins were inserted upstream of aph(3')-III resulting in a stable construct of 6174 bp that was used to transform both E. coli and Campylobacter.

Publication types

  • Comparative Study

MeSH terms

  • Base Sequence
  • Blotting, Southern
  • Campylobacter jejuni / genetics*
  • Chromosome Mapping
  • DNA Primers
  • Escherichia coli / genetics
  • Genetic Vectors / genetics*
  • Kanamycin Resistance / genetics
  • Molecular Sequence Data
  • Plasmids / genetics*
  • Sequence Analysis, DNA
  • Transformation, Bacterial

Substances

  • DNA Primers