title: Detection of rpoB gene mutations using helicase-dependent amplification.
abstract: For patients infected with tuberculosis, detection of rpoB gene mutations is critical for diagnosing drug-resistant strains of the causative pathogen, Mycobacterium tuberculosis (MTB). Traditional approaches to drug resistance include culture, which is very slow. Recently described real-time polymerase chain reaction approaches have addressed turnaround time but at relatively high cost. Here, we describe a novel amplification method, termed blocked-primer helicase-dependent amplification, for amplifying rpoB gene sequences in MTB. Resultant amplicon is hybridized to a probe set arrayed on a modified silicon-based chip to determine if there is any mutation in that region. Using this method, we could detect the majority of clinically relevant mutations in rpoB gene.
authors.affiliation: Great Basin Corporation, Salt Lake City, UT, USA.
authors.forename: Wanyuan
authors.initials: W
authors.lastname: Ao
authors.forename: Robert
authors.initials: R
authors.lastname: Jenison
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D004265:DNA Helicases / Q000737:chemistry; D004252:DNA Mutational Analysis / Q000379:methods*; D017931:DNA Primers / Q000235:genetics; D012321:DNA-Directed RNA Polymerases; D009169:Mycobacterium tuberculosis / Q000235:genetics; D021141:Nucleic Acid Amplification Techniques; D009693:Nucleic Acid Hybridization
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; C505805:rpoB protein, Mycobacterium tuberculosis; D012321:DNA-Directed RNA Polymerases; D004265:DNA Helicases