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Digoxigenin‐labelled peptide nucleic acid to detect lactobacilli PCR amplicons immobilized on membranes from denaturing gradient gel electrophoresis
Author(s) -
Burton J.P.,
McCormick J.K.,
Cadieux P.A.,
Reid G.
Publication year - 2003
Publication title -
letters in applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.698
H-Index - 110
eISSN - 1472-765X
pISSN - 0266-8254
DOI - 10.1046/j.1472-765x.2003.01281.x
Subject(s) - temperature gradient gel electrophoresis , amplicon , peptide nucleic acid , digoxigenin , biology , microbiology and biotechnology , nucleic acid , gel electrophoresis , lactobacillus , southern blot , 16s ribosomal rna , nucleic acid thermodynamics , hybridization probe , dna , biochemistry , bacteria , polymerase chain reaction , gene , rna , gene expression , genetics , in situ hybridization
Aims: To develop a digoxigenin (DIG)‐labeled peptide nucleic acid (PNA) probe for the detection of Lactobacillus ‐related genera amongst eubacterial amplicons obtained from vaginal samples using denaturing gradient gel electrophoresis (DGGE) blots. Methods and Results: Part of the 16S rRNA gene sequence was used as a target for the PNA probe. After confirming probe specificity using chromosomal DNA from species and isolates that have been detected in the urogenital tract, it was successfully used to detect lactobacilli amplicons generated using eubacterial‐specific 16S rRNA gene‐targeted primers from vaginal tract samples immobilized on membranes from DGGE. Conclusions: The Lactobacillus ‐specific PNA probe could distinguish between DNA fragments from lactobacilli in a DGGE gel from other bacterial species, including those that migrated to a similar position. Significance and Impact of the Study: The use of the DIG‐labelled PNA probe on blots of eubacterial PCR products from DGGE gels can be used to specifically detect lactobacilli in complex vaginal samples.