Gene replacement in Lactobacillus helveticus.
An efficient method for gene replacement in Lactobacillus helveticus CNRZ32 was developed by utilizing pSA3 as an integration vector. This plasmid is stably maintained in CNRZ32 at 37 degrees C but is unstable at 45 degrees C. This method consisted of a two-step gene-targeting technique: (i) chromosomal integration of a plasmid carrying an internal deletion in the gene of interest via homologous recombination and (ii) excision of the vector and the wild-type gene via homologous recombination, resulting in gene replacement. By using this procedure, the chromosomal X-prolyl dipeptidyl aminopeptidase gene (pepXP) of CNRZ32 was successfully inactivated.
Bhowmik
T
T
Department of Food Science, University of Wisconsin-Madison 53706.
Fernández
L
L
Steele
J L
JL
D002876:Chromosomes, Bacterial; D018819:Dipeptidyl Peptidase 4; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000235:genetics; D018274:Electroporation; D017353:Gene Deletion; D018014:Gene Transfer Techniques; D005798:Genes, Bacterial; D005821:Genetic Techniques; D005822:Genetic Vectors; D007778:Lactobacillus / Q000235:genetics* / Q000254:growth & development; D010957:Plasmids; D011995:Recombination, Genetic; D015183:Restriction Mapping
D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; D018819:Dipeptidyl Peptidase 4