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Applied and Environmental Microbiology, May 2002, p. 2209-2213, Vol. 68, No. 5
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.5.2209-2213.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Rapid PCR-Based Method Which Can Determine Both Phenotype and Genotype of Lactococcus lactis Subspecies

Masaru Nomura,* Miho Kobayashi, and Takashi Okamoto

Department of Animal Products Research, National Institute of Livestock and Grassland Science, Norindanchi, Tsukuba 305-0901, Japan

Received 27 August 2001/ Accepted 15 February 2002

A highly efficient, rapid, and reliable PCR-based method for distinguishing Lactococcus lactis subspecies (L. lactis subsp. lactis and L. lactis subsp. cremoris) is described. Primers complementary to positions in the glutamate decarboxylase gene have been constructed. PCR analysis with extracted DNA or with cells of different L. lactis strains resulted in specific fragments. The length polymorphism of the PCR fragments allowed a clear distinction of the L. lactis subspecies. The amplified fragment length polymorphism with the primers and the restriction fragment length polymorphism of the amplified products agreed perfectly with the identification based on genotypic and phenotypic analyses, respectively. Isolates from cheese starters were investigated by this method, and amplified fragments of genetic variants were found to be approximately 40 bp shorter than the typical L. lactis subsp. cremoris fragments.

* Corresponding author. Mailing address: Department of Animal Products Research, National Institute of Livestock and Grassland Science, Norindanchi, Tsukuba 305-0901, Japan. Phone: 81 298 38 8688. Fax: 81 298 38 8683. E-mail: nomura{at}affrc.go.jp.

Applied and Environmental Microbiology, May 2002, p. 2209-2213, Vol. 68, No. 5
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.5.2209-2213.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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