Replication and inheritance ofNocardiaplasmid pC1

Abstract

Nocardia sp. C-14-1, isolated from acrylic fiber wastewater, can degrade long-chain alkanes and succinonitrile efficiently. Here we report the characterization of an indigenous plasmid pC1. The overall nucleotide sequence of pC1 consisted of 5841 bp. The five ORFs, encoding a DNA recombinase, replication protein (Rep(pC1)) and three proteins of unknown function, were predicted on pC1. The Rep(pC1) displayed its homology with the Rep of Rhodococcus large plasmid p33701, suggesting a theta type of replication. An Escherichia coli plasmid (containing the single rep(pC1) gene) propagated autonomously in low copy number in Nocardia or Rhodococcus, suggesting that rep(pC1) was an essential gene for plasmid replication. The plasmid (containing the single rep(pC1) gene) presented as inheritance unstable hints that other pC1 loci were required for the stable inheritance of plasmids. By comparison of the plasmid-borne Rep proteins, we classify Rhodococcus or Nocardia plasmids into four groups.

Development of a vector and host system and characterization of replication of plasmid pSQ10 in moderately halophilic <italic>Nocardiopsis</italic>

“…Electroporation of Rhodococcus sp. N1037 followed Shen et al [19]. Escherichia coli strains DH5a [supE44, DlacU169 (f80lacZDM15), hsdR17, recA1, endA1, gyrA96, thi-1, relA1; Life Technologies, Carlsbad, USA] and ET12567 (dam dcm hsdM cm kan) was used as cloning hosts.…”

Construction of novel shuttle expression vectors for gene expression in <i>Bacillus subtilis</i> and <i>Bacillus pumilus</i>

“…Determination of the plasmid copy number. The copy number of the plasmid in the cells was determined according to the method described by Shen et al (2006). Briefly, B. subtilis S1-4 and B. subtilis WB600 harboring pSU03-AP were grown at 37°C for 24 h in 50 ml LB broth without or with 50 µg/ml kanamycin, respectively.…”