Clotblood dna kit

Genomic DNA was extracted from peripheral blood lymphocytes using the ClotBlood DNA Kit (Cwbio, Beijing) and detected using an ultraviolet spectrophotometer (Beckman, USA), OD260/OD280: 1.6–1.9.
We selected three tagSNPs located in the fibrinogen alpha chain gene (FGA) by Haploview 4.2 using genotype data from Genome Browser release #27 in the HapMap Project (http://www.hapmap.org). FGA is mapped to Chromosome 4q28 and contains six exons. These three tagSNPs included rs2070011 (in the 5′UTR of FGA), rs2070016 (in intron 4 of FGA), and rs2070022 (in the 3′UTR of FGA). The primers for genotyping these three tagSNPs were designed using the AssayDesigner3.1 and are listed in Table 1.
SNP genotyping was determined by the allele-specific matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF-MS) using the Sequenom MassARRAY platform in Bio-X Institutes, Beijing [6 ]. Using a MassARRAY Nanodispenser (Sequenom, San Diego, CA, USA), standardized genotyping reactions were dispensed onto a 384-well spectroCHIP [7 (link)]. The repeated control samples were set in every genotyping plate and the concordance was more than 99%.

We collected 5 mL blood sample from every patient and stored the specimens at −20°C in nonanticoagulant, plexiglass tubes. Genomic DNA was extracted from peripheral blood lymphocytes by using a commercial DNA extraction kit (ClotBlood DNA kit, Cwbio, Beijing, China), according to the manufacturer's instructions. DNA concentration and purity were detected by ultraviolet spectrophotometer (Beckman, USA) with OD₂₆₀/OD₂₈₀ between 1.6 and 1.9 which we thought to be satisfactory.

5 ml of peripheral blood obtained from each participant was stored in EDTA-containing tubes. Genomic DNA was extracted according to the instruction of the ClotBlood DNA Kit (Cwbio, Beijing, China). The DNA concentration was measured using the ultraviolet spectrophotometer (Beckman, USA). Genotypes were detected by iPLEX Gold reagent set (Sequenom, San Diego, CA, USA). The standard polymerase chain reaction (PCR) was performed and PCR products were assayed by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS). The PCR primers were designed by Assay Designer 3.1 (Table 1). Samples that failed genotyping were removed from the analyses. The internal blinded quality control of the samples showed 97% consistency.