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  • Title: An efficient PCR--SSCP-based method for detection of a chloroquine resistance marker in the PfCRT gene of Plasmodium falciparum.
    Author: Mishra S, Raj DK, Hazra RK, Dash AP, Supakar PC.
    Journal: Trans R Soc Trop Med Hyg; 2006 Mar; 100(3):243-7. PubMed ID: 16288790.
    Abstract:
    The spread of chloroquine resistance throughout the world poses a major problem in combating malaria. In the present study, an efficient polymerase chain reaction-single strand conformational polymorphism (PCR--SSCP)-based assay detected the PfCRT K76T point mutation, which is a marker for chloroquine resistance. For the first time, we have used a PCR--SSCP-based technique to identify the mutation in a single-step labelling reaction during PCR and SSCP gel electrophoresis. This assay is 100% efficient, giving no false-positive or -negative results, and can be carried out within a short bench time. We have successfully analysed 120 natural isolates using the PCR-SSCP method for detection of the chloroquine resistance marker and found 91 of the 120 samples to show the PfCRT T76 mutation, and 71% (65 of the 91 samples) showed a positive correlation with chloroquine resistance from the clinical data of the patients. The PCR-SSCP technique can also be applied for the detection of new haplotypes of the PfCRT gene and surveillance of chloroquine-resistant malaria in malaria-endemic localities around the world.
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