Detection of rpoB mutations in rifampin resistant Mycobacterium tuberculosis strains by the reverse dot blot hybridization method
- VernacularTitle:应用反向斑点杂交法检测耐利福平结核分支杆菌rpoB基因突变
- Author:
Maoyin PANG
;
Wenhong ZHANG
;
Zhongyi HU
;
Al ET
;
- Publication Type:Journal Article
- Keywords:
Reverse dot blot hybridization;
Drug Tolerance;
Mycobacterium tuberculosis;
Mutation;
Rifampin
- From:
Chinese Journal of Infectious Diseases
1997;0(04):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To develop new method for rapid detection of mutations in rpoB gene related with resistance to rifampin of M. tuberculosis . Methods According to the sequence of wild type M. tuberculosis, five oligonucleotide probes covering the 69 bp hyper variable region of rpoB gene were designed and immobilized on nylon membrane strips. Thereafter, the target rpoB gene fragment was obtained by PCR using biotin labeled primers and thereafter the PCR product was denatured and hybridized with probes on membrane. The results of reverse dot blot hybridization were compared with the results of drug sensitivity test and sequencing. Results PCR products from 36 RFP resistant and 22 RFP susceptible isolates were detected by the assay of reverse dot blot hybridization, showing that the susceptibility and the specificity rate were 88.9% and 86.4% respectively,and the coincidence with biochemical method and sequencing is 87.9% and 89.7% respectively. Conclusions Reverse dot blot hybridization is a rapid and sensitive method to detect the rpoB gene mutations,which may be used in early detection of the resistance of M. tuberculosis to rifampin.
