Specific detection of bacillus anthracis using a TaqMan mismatch amplification mutation assay.
Single nucleotide polymorphisms (SNPs) are increasingly recognized as important diagnostic markers for the detection and differentiation of Bacillus anthracis. The use of SNP markers for identifying B. anthracis DNA in environmental samples containing genetically similar bacteria requires the ability to amplify and detect DNA with single nucleotide specificity. We designed a TaqMan mismatch amplification mutation assay (TaqMAMA) around a SNP in the plcR gene of B. anthracis. The assay permits specific, low-level detection (25 fg DNA) of this B. anthracis-specific SNP, even in the presence of environmental DNA extracts containing a 20,000-fold excess of the alternate allele. We anticipate that the ability to selectively amplify and detect low copy number DNAs with single nucleotide specificity will represent a valuable tool in the arena of biodefense and microbial forensics.
Easterday
William R
WR
Northern Arizona University, Flagstaff, AZ 86011-5640, USA.
Van Ert
Matthew N
MN
Zanecki
Shaylan
S
Keim
Paul
P
D001408:Bacillus anthracis / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification*; D020137:Base Pair Mismatch / Q000235:genetics; D004252:DNA Mutational Analysis / Q000379:methods*; D017403:In Situ Hybridization / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D016415:Sequence Alignment / Q000379:methods*; D017422:Sequence Analysis, DNA / Q000379:methods*; D019914:Taq Polymerase / Q000378:metabolism
D019914:Taq Polymerase