%0 Journal Article
%T Plasmid-based controls to detect rpoB mutations in Mycobacterium tuberculosis by quantitative polymerase chain reaction-high-resolution melting
%A Silva
%A Joas Lucas da
%A Leite
%A Gabriela Guimaraes Sousa
%A Bastos
%A Gisele Medeiros
%A Lucas
%A Beatriz Cacciacarro
%A Shinohara
%A Daniel Keniti
%A Takinami
%A Joice Sayuri
%A Miyata
%A Marcelo
%A Fajardo
%A Cristina Moreno
%A Luchessi
%A Andr¨¦ Ducati
%A Leite
%A Clarice Queico Fujimura
%A Cardoso
%A Rosilene Fressatti
%A Hirata
%A Rosario Dominguez Crespo
%A Hirata
%A Mario Hiroyuki
%J Mem¨®rias do Instituto Oswaldo Cruz
%D 2013
%I Instituto Oswaldo Cruz, Minist¨¦rio da Sa¨²de
%R 10.1590/S0074-02762013000100017
%X quantitative polymerase chain reaction-high-resolution melting (qpcr-hrm) analysis was used to screen for mutations related to drug resistance in mycobacterium tuberculosis. we detected the c526t and c531t mutations in the rifampicin resistance-determining region (rrdr) of the rpob gene with qpcr-hrm using plasmid-based controls. a segment of the rrdr region from m. tuberculosis h37rv and from strains carrying c531t or c526t mutations in the rpob were cloned into pgem-t vector and these vectors were used as controls in the qpcr-hrm analysis of 54 m. tuberculosis strains. the results were confirmed by dna sequencing and showed that recombinant plasmids can replace genomic dna as controls in the qpcr-hrm assay. plasmids can be handled outside of biosafety level 3 facilities, reducing the risk of contamination and the cost of the assay. plasmids have a high stability, are normally maintained in escherichia coli and can be extracted in large amounts.
%K drug resistance
%K rifampicin
%K mycobacterium tuberculosis.
%U http://www.scielo.br/scielo.php?script=sci_abstract&pid=S0074-02762013000100017&lng=en&nrm=iso&tlng=en