New multiplex allele-specific PCR assay targeting katG codon 315 variation for detection of isoniazid resistant Mycobacterium tuberculosis strains / 中华检验医学杂志

ABSTRACT

Objective To develop a new multiplex allele-specific PCR(MAS-PCR) assay to detect mutation in codon 315 of katG gene of Mycobacterium tuberculosis , mutation in this codon has been reported to be able to account for the Mycobacterium tuberculosis clinical isolates resistant to isoniazid(INH).Method Based on the sequence of katG gene of Mycobacterium tuberculosis , three specific primers are designed to carry out the MAS-PCR, 84 purified DNA preparation with known katG 315 variation detected by PCR-restriction fragment length polymorphism (PCR-RFLP) are used to optimize PCR.Results 84 Mycobacterium tuberculosis clinical stains are detected by the MAS-PCR and PCR-RFLP, respectively.The sensitivity of detection by MAS-PCR is 77.8%,and the specificity is 95.2%.katG mutation S315N(AGC→AAC), neglected in RFLP, can be detected by MAS-PCR.Conclusion MAS-PCR assay is sensitive, specific, economic and easy to carry out , can be used in clinical laboratories to detect the INH-resistant Mycobacterium tuberculosis strains.