Testing of a whole genome PCR scanning approach to identify genomic variability in four different species of lactic acid bacteria.
Genomes can be markedly heterogeneous in conspecific bacterial strains. Genome sequences can be used to analyze genome plasticity via a PCR(2) (plasticity of chromosome revealed by PCR) approach. Small-sized chromosomes can indeed be fully amplified by long-range PCR with a set of primers designed using a reference strain and then applied to several other strains. Analysis of the resulting patterns can reveal genome plasticity. GenoFrag, a software package for the design of primers optimized for PCR(2) [N. Ben Zakour, M. Gautier, R. Andonov, D. Lavenier, M.F. Cochet, P. Veber, A. Sorokin, Y. Le Loir, GenoFrag: Software to design primers optimized for whole genome scanning by long-range PCR amplification, Nucleic Acids Res. 32 (2004) 17-24] was developed for the analysis of bacterial genome plasticity by whole genome amplification in approximately 10-kb-long fragments. By applying GenoFrag, we provide herewith evidence that genome plasticity can be analyzed in lactic acid bacteria using a PCR(2) approach. The genome sequences of Lactococcus lactis IL1403, Lactobacillus plantarum WCFS1, Lactobacillus bulgaricus ATCC11842 and Bifidobacterium longum NCC2705 were used to design four sets of primers. Each set was evaluated in silico to check that it ensured optimum coverage of the bacterial chromosome. To validate the primers generated by GenoFrag, a subset of primers was successfully used in LR-PCR experiments on genomic DNA from four L. bulgaricus strains.
Ben Zakour
Nouri
N
Laboratoire de Microbiologie, UMR1253, Institut National de la Recherche Agronomique, Agrocampus, CS84215, 65, rue de Saint Brieuc, 35042 Rennes Cedex, France.
Grimaldi
Christine
C
Gautier
Michel
M
Langella
Philippe
P
Azevedo
Vasco
V
Maguin
Emmanuelle
E
Le Loir
Yves
Y
D015373:Bacterial Typing Techniques; D001644:Bifidobacterium / Q000235:genetics*; D017931:DNA Primers; D004269:DNA, Bacterial / Q000235:genetics; D014644:Genetic Variation; D016680:Genome, Bacterial; D019344:Lactic Acid / Q000378:metabolism; D007778:Lactobacillus / Q000235:genetics*; D048191:Lactiplantibacillus plantarum / Q000235:genetics; D013294:Lactococcus lactis / Q000235:genetics; D016133:Polymerase Chain Reaction; D012984:Software
D017931:DNA Primers; D004269:DNA, Bacterial; D019344:Lactic Acid