RAPID AND CLEAR DETECTION OF ABO GENOTYPES BY SIMULTANEOUS PCR-RFLP METHOD

We reported a new approach of ABO genotyping by a polymerase chain rea ction and restriction fragment length polymorphism method. Instead of amplifying the loci containing the positions of nucleotides 258 and 70 0 of cDNA of the A transferase separately, we successfully amplified t hese 2 loci together in one reaction mixture using 2 sets of primers. The amplified DNA products were digested at the same time with restric tion enzymes Kpn I and Alu I. The digested DNA products were then sepa rated by electrophoresis on polyacrylamide gel. In addition, we evalua ted the influence of various amplification parameters (concentration o f template DNA, primers, Tag DNA polymerase, MgCl2, and number of cycl es). In particular, high Mg2+ concentration (3.5 mM) made effective am plification of this locus without producing any unspecific band. By us ing that optimized condition for PCR, together with a simultaneous app roach, our study proved to be time saving, more economic, and convenie nt in interpreting the results.