Evaluation of Five Methods Used to Extract Deoxyribonucleic Acid (DNA) From Human Malaria Parasitized Blood Spotted on the Filter Paper
Keywords:
malaria, blood spot, extraction, purity, PCRAbstract
Efficient deoxyribonucleic acid (DNA) extraction is critical for a good polymerase chain reaction (PCR) performance in molecular diagnostic, genomic and epidemiological studies of malaria. Blood collected on filter paper is a pratical method for this puposes in high parasitemic individuals. In this paper we evaluated five different methods (Commercial kit, Chelex-saponin, microwave, methanol and Tris-EDTA) for DNA extraction. Blood specimens from six asymptomatic Plasmodium sp.-infected patients in Jayapura General Hospital were spotted onto filter paper, dried, and transported for processing. Evaluation of the results of DNA extraction was carried out by measuring yields using spectrophotometry and PCR amplification for mitochondrial gene. Results showed that the best results of PCR obtanied only with commercial kit method, whereas very low quality of PCR by microwave and no amplified DNA for three other methods. The purity of DNA for all methods was low. The sensitivity of detection for kit was approximately 100% for P. falciparum, P. malariae and P. vivax, whereas microwave method, even though resulted in high concentration of DNA, did not or slightly show the effectiveness in DNA amplification that may due to high inhibitor contents. Even though chelex method was widely used by many malaria laboratories, in this experiment we did not found its effectiveness. We concluded that commercial extraction kit method is well-suited for the DNA extraction in PCR-based assays of malaria.
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