Rapid PCR-based procedure to identify lactic acid bacteria: application to six common Lactobacillus species.
The goal of this study was to develop a method allowing rapid identification of the lactic acid bacteria strains in use in the laboratory (Lactobacillus plantarum NCIMB8826; L. fermentum KLD; L. reuteri 100-23; L. salivarius UCC43321; L. paracasei LbTGS1.4; L. casei ATCC393), based on PCR amplification of 16S RNA coding sequences. First, specific forward oligonucleotides were designed in the variable regions of 16S RNA coding sequences of six Lactobacillus strains. The reverse oligonucleotide was designed in the region where the sequences were homologous for the six strains. The expected size of the amplification product was +/-1000 bp. The specificity of the method was tested on total chromosomal DNA. For five out of the six strains, the amplification of the fragment was strain-specific and the method was directly applicable to colonies. For the strain L. casei ATCC393, an additional argument to the classification of this bacteria in the paracasei group could be proposed. Validation of the developed method was performed by applying it to six Lactobacillus reference strains and to various species of bacteria.
Chagnaud
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P
Département de Microbiologie des Ecosystèmes, Institut Pasteur de Lille; 1, rue du Professeur Calmette, BP 245, 59019 Cedex, Lille, France. patrice.chagnaud@pasteur-lille.fr
Machinis
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K
Coutte
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LA
Marecat
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A
Mercenier
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D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics; D004275:DNA, Ribosomal / Q000032:analysis / Q000235:genetics; D006801:Humans; D007778:Lactobacillus / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012015:Reference Standards; D021521:Ribotyping; D013045:Species Specificity
D017931:DNA Primers; D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S