Temperature-mediated heteroduplex analysis for the detection of drug-resistant gene mutations in clinical isolates of Mycobacterium tuberculosis by denaturing HPLC, SURVEYOR nuclease.

Abstract:

:Denaturing high-performance liquid chromatography (DHPLC) is a relatively new technique, which utilizes heteroduplex formation between wild-type and mutated DNA strands to identify point mutations. Heteroduplex molecules are separated from homoduplex molecules by ion-pair, reverse-phase liquid chromatography on a special column matrix with partial heat denaturation of the DNA strands. In order to investigate the application of this method for point mutation detection in drug-resistant genes of Mycobacterium tuberculosis, katG, rpoB, embB, gyrA, pncA and rpsL genes, which are responsible for isoniazid, rifampicin, ethambutol, fluoroquinolone, pyrazinamide and streptomycin resistance, respectively, were detected by temperature-mediated DHPLC in 10 multidrug-resistant and 10 drug-susceptible clinical isolates. The DHPLC data were compared with those from a conventional MIC test. The results show that DHPLC is cost-effective with high capacity and accuracy, and is potentially useful for genotypic screening for mutations associated with anti-tuberculosis drug resistance.

journal_name

Microbes Infect

journal_title

Microbes and infection

authors

Shi R,Otomo K,Yamada H,Tatsumi T,Sugawara I

doi

10.1016/j.micinf.2005.06.008

subject

Has Abstract

pub_date

2006-01-01 00:00:00

pages

128-35

issue

1

eissn

1286-4579

issn

1769-714X

pii

S1286-4579(05)00234-0

journal_volume

8

pub_type

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