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Figure 1. GSTT1 present and null alleles. bp: Base pair.
(Figure omitted. See article PDF.)
Figure 2. Gel showing the genotype for GSTT1 null/present polymorphism. Lane M contains a 100 bp DNA ladder, lane 1 the null homozygote (NN ), lane 2 heterozygote (PN ) and lane 3 the present homozygote (PP ). bp: Base pair.
(Figure omitted. See article PDF.)
Figure 3. Gel obtained for actual genotyping.
(Figure omitted. See article PDF.)
Glutathione S-transferase (GST) th[theta] is a cytosol protein with GST activity [1], which is one of the main processes of phase II detoxification of xenobiotic compounds, such as carcinogens included in cigarette smoke [2,3]. GSTT1 encoding GST th[theta] is located in 22q11.2. The null/present polymorphism has been studied for the risks of many diseases including cancers [4-6].
GSTT1 null/present genotyping has been conducted by the detection of only the present allele, resulting in the present type including a heterozygote (PN genotype) and the present homozygote (PP genotype). When the present allele is not detected, the DNA is therefore determined to be the null (NN ) type. Sprenger and colleagues invented a genotyping method to separate the PN and PP genotypes by using primers amplifying 1460 base pair (bp) fragments [7]. The method was adopted by another study [8], but its use was still limited, possibly as a result of the difficulty in conducting the long chain PCR for the samples derived from epidemiological studies.
This article reports a PCR method with a new set of primers for the GSTT1 present allele, which can be used with the null allele primers within one tube. The method was applied to samples taken from patients attending a healthcare check-up in Japan, which demonstrates the applicability of the method for epidemiological studies.
GSTT1 DNA sequence
As Sprenger and colleagues described, the GSTT1 -null genotype lacks 5022 bp including the GSTT1 gene [7]. The connection part of the GSTT1- null allele has an identical common 403-bp sequence to that in HA5 and HA3 genes flanking the GSTT1 gene. The left side of the common sequence of the null allele is the same as the corresponding sequence of HA5 and the right side of HA3 , as demonstrated in Figure 1.
Primers...