Abi prism 7900 sequence detection system software

Venous blood samples from all participants were obtained using 20 g/L ethylene diamine tetra-acetic acid anticoagulant. Genomic DNA was extracted from 250 µl of venous blood using a QIAamp DNA Blood Mini Kit (Qiagen, Valencia, CA, USA) and stored at –80℃ until analysis according to the manufacturer’s instructions. The TGF-β1 SNPs rs1800469 and rs1982073 were genotyped using the TaqMan SNP Genotyping Assay (Applied Biosystems, Foster City, CA, USA). We performed PCR amplification^{22 (link)} with 100 ng of genomic DNA, 0.1 µM of each probe, 0.2 µM of each primer, 200 µM of each deoxyribonucleotide, 3 mM MgCl₂, and 1 U Platinum Taq DNA polymerase. PCR amplification was carried with an initial denaturation at 95℃ for 2 min, followed by 40 cycles of denaturation at 95℃ for 15 s and annealing at 65℃ for 30 s using the 7900HT Fast Real-Time PCR System (Applied Biosystems).^{23 (link)} ABI PRISM 7900 Sequence Detection System software version 2.3 (Applied Biosystems) was used to perform data analyses. For confirmation, approximately 5% of the samples were selected randomly for repeated genotyping.