Abstract: According to the fact that Nisin-resistant gene and lactose-fermenting gene often have the same locus, five Nisin-resistant strains were screened from fresh milk with a selective medium (GM17) supplemented with 0.004 g/100 mL bromocresol purple and 300 IU/mL Nisin. By using skimmed culture medium, Gram s staining and microscopic examination, five strains were identified as Lactococcus lactis. PCR amplification of Nisin-resistant gene (NSR) was carried out with chromosome and plasmid DNA as templates. An expected PCR product with 400 bp was achieved from three out of five strains. The 16S rDNA sequence analysis revealed that three strains named as N1, N2 and N3 belonged to Lactococcus lactis. The full-length of Nisin-resistant determinant gene (NSR) with 1 kb in size was cloned with the plasmids N1, N2 and N3 as templates, respectively. The PCR product of N1 was sequenced and the amplicon exhibited 99% similarity with the published sequence and was therefore confirmed as NSR by Blast analysis.

Key words: Nisin-resistant strain, screening, Nisin-resistant gene (NSR), cloning