Duplex polymerase chain reaction with confronting two-pair primers (PCR-CTPP) for genotyping alcohol dehydrogenase beta subunit (ADH2) and aldehyde dehydrogenase 2 (ALDH2).
OBJECTIVE
Alcohol dehydrogenase beta subunit (ADH2) Arg47His and aldehyde dehydrogenase 2 (ALDH2) Glu487Lys were genotyped by a duplex polymerase chain reaction (PCR) with confronting two-pair primers (PCR-CTPP), which allows DNA amplification with one-tube PCR including eight primers, and subsequent electrophoresis.


METHODS
Several PCR conditions were tested to establish the optimal conditions for distinguishing the allele-specific bands for the two polymorphisms. Under the optimal PCR conditions, 454 Japanese health check-up examinees were genotyped.


RESULTS
The allele-specific bands were successfully amplified under the optimal conditions of the duplex PCR-CTPP. The genotype distributions were within the Hardy-Weinberg equilibrium. The bands produced by the duplex PCR-CTPP genotyping were clearer than those produced by PCR-CTPP, conducted solely for ADH2.


CONCLUSIONS
ADH2 Arg47His and ALDH2 Glu487Lys were successfully genotyped by this newly developed duplex PCR-CTPP, an inexpensive and time-saving genotyping tool, which will be useful in epidemiological studies on alcoholism, as well as risk estimation of alcohol-related diseases.
Tamakoshi
Akiko
A
Department of Preventive Medicine/Biostatistics and Medical Decision Making, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550, Japan.
Hamajima
Nobuyuki
N
Kawase
Haruya
H
Wakai
Kenji
K
Katsuda
Nobuyuki
N
Saito
Toshiko
T
Ito
Hidemi
H
Hirose
Kaoru
K
Takezaki
Toshiro
T
Tajima
Kazuo
K
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000426:Alcohol Dehydrogenase / Q000235:genetics*; D000444:Aldehyde Dehydrogenase / Q000235:genetics*; D000071396:Aldehyde Dehydrogenase, Mitochondrial; D016009:Chi-Square Distribution; D017931:DNA Primers / Q000235:genetics*; D005260:Female; D005838:Genotype; D006801:Humans; D008297:Male; D008875:Middle Aged; D009692:Nucleic Acid Heteroduplexes / Q000737:chemistry* / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*
D017931:DNA Primers; D009692:Nucleic Acid Heteroduplexes; D000426:Alcohol Dehydrogenase; C096127:ALDH2 protein, human; D000444:Aldehyde Dehydrogenase; D000071396:Aldehyde Dehydrogenase, Mitochondrial