Highly Specific Culture-Independent Detection of YGNGV Motif-Containing Pediocin-Producing Strains

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• 作者：S. K. Sood (1)
  B. Vijay Simha (1)
  Rashmi Kumariya (1)
  Anita Kumari Garsa (1)
  Jitender Mehla (1)
  Sunita Meena (1)
  Puja Lather (1)
  
• 关键词：Pediocin ; YGNGV motif ; PCR ; based detection ; Pediococcus ; Enterococcus ; Lactobacillus ; Lactic acid bacteria
• 刊名：Probiotics and Antimicrobial Proteins
• 出版年：2013
• 出版时间：March 2013
• 年：2013
• 卷：5
• 期：1
• 页码：37-42
• 全文大小：480KB
• 参考文献：1. Abriouel H, Franz CMAP, Omar NB, Galvez A (2011) Diversity and applications of / Bacillus bacteriocins. FEMS Microbiol Rev 35:201-32 CrossRef
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• 作者单位：S. K. Sood (1)
  B. Vijay Simha (1)
  Rashmi Kumariya (1)
  Anita Kumari Garsa (1)
  Jitender Mehla (1)
  Sunita Meena (1)
  Puja Lather (1)
  
  1. Division of Animal Biochemistry, National Dairy Research Institute, Karnal, 132001, Haryana, India
  
• ISSN：1867-1314

文摘

A novel method based on (1) initial microbiological screening and (2) a highly specific PCR is described for selection of strains expressing YGNGV motif-containing pediocin. Initial screening is carried out using spot on the lawn assay for selection of acid-free, hydrogen peroxide (H2O2)-free and secreted heat-stable inhibitory activity producing strains. This is followed by highly specific PCR for amplification of 406-bp fragment using forward primer: 5-tggccaatatcattggtggt-3-targeting signal peptide sequence of pediocin structural gene and reverse primer: 5-ctactaacgcttggctggca-3-encoding N-terminus of immunity gene. The assay was validated with Pediococcus pentosaceus NCDC273 and Pediococcus acidilactici NCDC252 using (1) digestion of amplified 406-bp fragment with HindIII restriction enzyme-producing two restriction fragments of expected sizes (227 and 179?bp), (2) nucleotide sequencing of 406-bp fragment from both strains found these pediocins identical to pediocin PA-1/AcH and (3) identification of both pediocins as pediocin PA-1 at protein level using RP-HPLC. The assay was used for screening six strains (3 pediococci, 2 lactobacilli and an Enterococcus faecium) producing acid-free, hydrogen peroxide (H2O2)-free and secreted heat-stable inhibitory activity. This resulted in the detection of three new strains (P. pentosaceus NCDC35, E. faecium NCDC124 and Lactobacillus plantarum NCDC20) producing YGNGV motif-containing pediocins.