The invention relates to gene detection of disease pathogens and particularly relates to a nucleic acid membrane strip and a kit for drug-resistant mutant gene detection of mycobacterium tuberculosis. The kit comprises the nucleic acid membrane strip which comprises a substrate and a nucleic acid probe fixedly arranged on the substrate; the nucleic acid probe comprises probes with the base sequences as shown in SEQ ID NO: 1-26. The kit for drug-resistant mutant gene detection of mycobacterium tuberculosis can be used for comprehensively and quickly detecting the drug-resistant mutant gene of mycobacterium tuberculosis, can be used for detecting the drug resistance of five main antituberculosis drugs in one test and can be used for quickly and immediately determining drug-resistant information of mycobacterium tuberculosis on the main antituberculosis drugs, so that prophylactic treatment and diagnosis are favorably performed clinically in advance, thereby alleviating the burden of a patient.