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W52546518 abstract "The emergence of drug-resistant strains of Mycobacterium tuberculosis is a serious public health problem. Many of the specific gene mutations that cause drug resistance in M. tuberculosis are point mutations. We are developing a PCR-peptide nucleic acid (PNA)-based ELISA as a diagnostic method to recognize point mutations in genes associated with isoniazid and rifampin resistance in M. tuberculosis. Specific point mutation-containing sequences and wild-type sequences of cloned mycobacterial genes were PCR-amplified, denatured, and hybridized with PNA probes bound to microplate wells. Using 15-base PNA probes, we established the hybridization temperatures (50°C–55°C) and other experimental conditions suitable for detecting clinically relevant point mutations in the katG and rpoB genes. Hybridization of PCR-amplified sequences that contained these point mutations with complementary mutation-specific PNAs resulted in significant increases in ELISA response compared with hybridization using wild-type-specific PNAs. Conversely, PCR-amplified wild-type sequences hybridized much more efficiently with wild-type PNAs than with the mutation-specific PNAs. Using the M. tuberculosis cloned genes and PCR-PNA-ELISA format developed here, M. tuberculosis sequences containing point mutations associated with drug resistance can be identified in less than 24 h." @default.
W52546518 created "2016-06-24" @default.
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W52546518 date "2002-03-01" @default.
W52546518 modified "2023-09-26" @default.
W52546518 title "Peptide Nucleic Acid Probe Detection of Mutations in Mycobacterium tuberculosis Genes Associated with Drug Resistance" @default.
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W52546518 doi "https://doi.org/10.2144/02323st01" @default.
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