A STRATEGY FOR RAPID DETECTION OF ISONIAZIDE RESISTANCE IN Mycobacterium tuberculosis BASED ON MULTIPLEX PCR USING LOCAL RTTH DNA POLYMERASE AND INTERPRETATION OF AMPLIFICATION PATTERNS

Abstract

This invention discloses a strategy for rapid detection of isoniazid resistance in Mycobacterium tuberculosis based on multiplex polymerase chain reaction (Multiplex PCR) using local rTth DNA polymerase and interpretation of amplification patterns. This strategy utilizes a specific primer combination that allows simultaneous amplification of the control fragment and the target fragment of the katG gene codon 315 mutation in a single PCR reaction. The amplification products are analyzed by 1% (w / v) agarose gel electrophoresis and interpreted based on the resulting DNA banding pattern to distinguish sensitive isolates, isolates carrying the katG315 mutation causing isoniazid resistance, and invalid test results. The use of local rTth DNA polymerase provides an alternative thermostable enzyme to support national diagnostic raw material independence without changing the detection principle.This strategy allows for rapid identification of katG315 mutations associated with isoniazid resistance without the need for DNA sequencing or further molecular analysis. The invention provides a simple, rapid, specific, easily interpretable, and cost-effective molecular diagnostic method to support the early detection of isoniazid-resistant tuberculosis.