New DNA Extraction Method for Diagnosis of Tuberculosis by Polymerase Chain Reaction / 대한임상병리학회지

ABSTRACT

BACKGROUND: To assess the clinical utility of new DNA extraction method, the authors attempted PCR using mycobacterial DNA extracted by Chelex 100 ion exchange resin method for 63 clinical samples in patients with pulmonary tuberculosis and compared with proteinase K method, simultaneously. METHODS: We used Chelex 100 ion exchange resin for preparation of DNA. Decontaminated sputums were mixed with resin and incubated at 56degrees C and 100degrees C without opening tube. After centrifugation, supernatants were used directly as template for PCR. 245 bps in primary PCR and 188 bps in nested PCR were amplified and analysed by agarose gel electrophoresis EtBr staining. RESULTS: Chelex 100 ion exchange resin method is more simple, rapid and reliable than proteinase K method, and during sample preparation, carry-over contamination loss of amplificated DNA, influence of organic solvents and cross-contamination are diminished. The results of PCR products are interpreted more distinctively in Chelex 100 ion exchange resin method than proteinase K method. CONCLUSIONS: In the basis of the results, it could be suggested that extraction of mycobacterial DNA by Chelex 100 ion exchange resin is more simple, rapid reliable method than that of conventional method for detection of mycobacterial DNA in patients with tuberculosis by polymerase chain reaction.