I am working on characterization of bacteriocin producing lactic acid bacteria from meat. I have identified the isolates on the basis of biochemical tests. Now I have to identify them on molecular basis using PCR. Can anyone suggest me the universal primers or any specific primer sequence for identification of lactobacilli and lactococci.
Hello Shilpshri,
here are the primers we use. These are universal bacterial primers. You can also use them for the identification by cycle sequencing:
91e: 5'- gga att caa a(gt) g aat tga cgg ggg c-3'
13b: 5'- cgg gat ccc agg ccc ggg aac gta ttc ac-3' (Amplification product size ~ 550bp)
Source: Frebourg: JCM 38:2 (2000)
PCR conditions: 5 min 95°C, (30s 95°C, 60s 55°C, 2 min 72°C) x 30, 5min 72°C, 4°C
Best wishes
Dirk
Dear Madam, Dubernet et al (2002) developed a polymerase chain reaction (PCR)-based method for the identification of lactobacilli at the genus level. One specific primer, LbLMA1-rev, was designed by analysing similarities between the nucleotide sequence of the spacer between the 16S and 23S rRNA genes in a number of Lactobacillus strains. Amplification with LbLMA1-rev and R16-1, a universal primer, generated a PCR product for 23 Lactobacillus species. Electrophoresis did not reveal any discrete bands when Escherichia coli, Lactococcus lactis, Leuconostoc mesenteroides, Streptococcus thermophilus, Carnobacterium pissicola, Pediococcus pentosaceus, Bifidobacterium bifidum,
Weissella confusa, Enterococcus hirae, Staphylococcus aureus or Listeria monocytogenes DNA were used as template.
FEMS Microbiology Letters 214 (2002) 271-275
Regarding Lactococci spp. :
Pu et al (2002) developed a PCR for the detection and identification of species and subspecies of the Gram-positive bacterial genus Lactococcus
Journal of Applied Microbiology, 93: 353–361. doi:10.1046/j.1365-2672.2002.01688.x
There is another work by Deasy et al (2000)
Systematic and Applied Microbiology Volume 23, Issue 4, December 2000, Pages 510-522. A Rapid PCR Based Method to Distinguish between Lactococcus and Enterococcus. But the paper is not free to download.
Thank you Dr. Dirk Schmidt. I will go for the primer sequence which you have suggested as per the reference of Frebourg: JCM 38:2 (2000).
Thanks to Dr. Rahul. I wll go through the references and try the technique in my research.
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