Journal Published Online: 01 November 1996
Volume 41, Issue 6

Rapid and Clear Detection of ABO Genotypes by Simultaneous PCR-RFLP Method

CODEN: JFSCAS

Abstract

We reported a new approach of ABO genotyping by a polymerase chain reaction and restriction fragment length polymorphism method. Instead of amplifying the loci containing the positions of nucleotides 258 and 700 of cDNA of the A transferase separately, we successfully amplified these 2 loci together in one reaction mixture using 2 sets of primers. The amplified DNA products were digested at the same time with restriction enzymes Kpn I and Alu I. The digested DNA products were then separated by electrophoresis on polyacrylamide gel. In addition, we evaluated the influence of various amplification parameters (concentration of template DNA, primers, Taq DNA polymerase, MgCl2, and number of cycles). In particular, high Mg2+ concentration (3.5 mM) made effective amplification of this locus without producing any unspecific band. By using that optimized condition for PCR, together with a simultaneous approach, our study proved to be time saving, more economic, and convenient in interpreting the results.

Author Information

Tun, Z
Osaka University Medical School, Osaka, Japan
Honda, K
Osaka University Medical School, Osaka, Japan
Nakatome, M
Osaka University Medical School, Osaka, Japan
Nasimul Islam, M
Osaka University Medical School, Osaka, Japan
Bai, H
Osaka University Medical School, Osaka, Japan
Ogura, Y
Osaka University Medical School, Osaka, Japan
Kuroki, H
Osaka University Medical School, Osaka, Japan
Yamazaki, M
Osaka University Medical School, Osaka, Japan
Terada, M
Osaka University Medical School, Osaka, Japan
Wakasugi, C
Osaka University Medical School, Osaka, Japan
Pages: 4
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Stock #: JFS14042J
ISSN: 0022-1198
DOI: 10.1520/JFS14042J