Abstract
A replication probe vector (pUB380) was constructed for detecting the replicon of LAB plasmid. A cryptic plasmid, pEV105, was isolated fromEnterococcus faeciumKLDS 6.0718. Multiple restriction endonuclease fragments of pEV105 were separately ligased to pUB380. After a series of subcloning and electrotransformation, the minimal replicon of pEV105 was isolated on a 2.5 kbPstI-XbalI fragment. Replicon based on this region followed a theta-type mechanism of replication inEnterococcus faeciumKLDS 6.0718. The minimal replicon DNA fragment was sequenced and the result shows that the gene sequence homology has 99.8 % compared with partial sequence of pCIZ2 fromEnterococcus faeciumL50. Five putative ORFs were concluded by software. The result of alignments indicated two ORFs, repAl05 and repBl05, encoding two putative proteins RepAl05 and RepBl05 of 245 and 178 amino acids respectively. The 2.5 kb fragment of minimal replicon as a stable replicon is feasible for constructing food-grade cloning and expressing vector of lactic acid bacteria.
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