A multiplex real-time PCR method for detection of GSTM1 and GSTT1 copy numbers.
OBJECTIVE
Deletion polymorphisms of Glutathione-S-transferase (GST) M1 and T1 are considered risk factors for various diseases. However, most previous studies only distinguished "null" and "non-null" genotypes. Our aim was to develop a reliable, high-throughput GSTM1/T1 genotyping method able to determine allele copy numbers.


METHODS
We developed a multiplex real time PCR method to distinguish between heterozygous (1/0) and homozygous (1/1) GSTM1 and GSTT1 genotypes. The principle of relative quantification was applied and an expectation-maximisation (EM) algorithm was developed to assign one of 3 possible genotypes: 1/1, 1/0 or 0/0 for each of the two genes.


RESULTS
1320 Caucasians were genotyped using the newly developed method. The observed genotype distributions did not deviate from the expected and were in Hardy-Weinberg equilibrium. GSTM1 duplication was detected in one sample.


CONCLUSIONS
This new semiquantitative genotyping method is a sensitive and promising tool for large-scale molecular epidemiological and clinical studies.
Timofeeva
Maria
M
German Cancer Research Center (DKFZ), Division of Epigenomics and Cancer Risk Factors (C010), Im Neuenheimer Feld 280, Heidelberg, Germany.
Jäger
Birgit
B
Rosenberger
Albert
A
Sauter
Wiebke
W
Wichmann
Heinz-Erich
HE
Bickeböller
Heike
H
Risch
Angela
A
D000465:Algorithms; D015139:Blotting, Southern; D004247:DNA / Q000032:analysis*; D017353:Gene Deletion; D018628:Gene Dosage; D020440:Gene Duplication; D005787:Gene Frequency; D005820:Genetic Testing / Q000379:methods; D005982:Glutathione Transferase / Q000032:analysis / Q000235:genetics*; D006801:Humans; D011110:Polymorphism, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D004247:DNA; D005982:Glutathione Transferase; C413545:glutathione S-transferase T1