Background: This study aims to investigate the usefulness of GeneXpert MTB/RIF (Xpert) in diagnosing rifampicin (RIF) resistance among the patients in the secluded sanatoria of North Korea. This article is a follow up of a report that explored the characteristics of drug resistance in North Korean Sanatoria, published in 2017. Xpert is used as a diagnostic tool to check for multidrug-resistant tuberculosis (MDR-TB) in specific areas of North Korea by the Eugene Bell Foundation (EBF). While there are other diagnostic tools are used to test for RIF resistance, there is no other feasible method to consistently test for MDR-TB in North Korea. Currently, EBF exclusively uses Xpert in tuberculosis and RIF resistance diagnosis when providing medical support for tuberculosis patients in North Korea, where there is limited international access. Methods: We performed Xpert using cultured tuberculous bacilli of North Koreans and compared the result to the corresponding phenotypic drug susceptibility tests (pDST) and rifampicin resistant-determining region sequencing (RRDR). All samples were target-sequenced at the rpoB region, where unique RIF resistance related mutations, potentially specific to North Korea, were discovered.Results: Using the RRDR sequencing as the gold standard, Xpert’s sensitivity and specificity was 94.9% and 88.0%, respectively. In addition, we discovered several rare or novel mutations among the North Korean Mycobacterium tuberculosis isolates.Conclusions: RIF detection in North Korea is possible using Xpert as an alternative to pDST, as this research has shown. This research is not only imperative to North Korea, but also in locations where there is limited access to conventional pDST.