Efficient gene inactivation in Bacillus anthracis.
A procedure for high-efficiency gene inactivation in Bacillus anthracis has been developed. It is based on a highly temperature-sensitive plasmid vector carrying kanamycin resistance cassette surrounded by DNA fragments flanking the desired insertion site. The approach was tested by constructing glutamate racemase E1 (racE1), glutamate racemase E2 (racE2) and comEC knock-out mutants of B. anthracis strain DeltaANR. Allelic replacements were observed at high frequencies, ranging from approximately 0.5% for racE2 up to 50% for racE1 and comEC. The system can be used for genetic validation of potential drug targets.
Shatalin
Konstantin Y
KY
Center for Pharmaceutical Biotechnology, University of Illinois, M/C 870, 900 S. Ashland Ave., Chicago, IL 60607, USA. shatalin@uic.edu
Neyfakh
Alex A
AA
D000591:Amino Acid Isomerases / Q000235:genetics; D001408:Bacillus anthracis / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D007613:Kanamycin Resistance / Q000235:genetics; D008780:Methyltransferases / Q000235:genetics; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000379:methods*; D010957:Plasmids / Q000235:genetics; D019998:Racemases and Epimerases / Q000235:genetics; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA
D001426:Bacterial Proteins; D004269:DNA, Bacterial; C111078:ErmTR protein, bacteria; D008780:Methyltransferases; D019998:Racemases and Epimerases; D000591:Amino Acid Isomerases; C062292:glutamate racemase