Sequenom massarray rs1000 system

Among the 11 SNPs selected, rs2048327 in SLC22A3 [13 (link)] and rs9349379 in PHACTR1 [26 (link)] are reportedly associated with CAD; other PHACTR1 and SLC22A3 gene SNPs were chosen at random [12 (link), 13 (link)]. Minor allele frequencies of all SNPs were >5% in the HapMap of the Chinese Han Beijing population. We used a GoldMag-Mini Whole Blood Genomic DNA Purification Kit (GoldMag Co. Ltd. Xi'an City, China) to extract DNA from whole blood. Extracted DNA was quantified using NanoDrop2000. Genotyping was done with the Sequenom MassARRAY RS1000 system using the standard protocol recommended by the manufacturer. The multiplexed SNP MassEXTENDED assay was designed using Sequenom Mass-ARRAY Assay Design 3.0 Software [27 ]. Data were managed and analyzed using Sequenom Typer 4.0 Software. Genotype was analyzed using a Sequenom MassARRAY RS1000 system and the standard protocol recommended by the manufacturer [27 , 28 (link)].

Among the 12 SNPs we selected, rs16944 in IL-1B [5 (link)] was chosen from previously published polymorphisms associated with AS, others were randomly chosen from the published genes (IL-1A and IL-1B) associated with AS. Minor allele frequencies of all SNPs were > 5%, in the HapMap of the Chinese Han CHB population. We adopted Gold Mag-Mini Whole Blood Genomic DNA Purification Kit (GoldMagCo. Ltd. Xi’an City, China) to extract DNA that was from whole blood. Quantification of the extracted DNA was performed using NanoDrop2000. Genotyping was done with the Sequenom MassARRAY RS1000 system using the standard protocol recommended by the manufacturer. The multiplexed SNP Mass EXTENDED assay was designed using Sequenom Mass-ARRAY Assay Design 3.0 Software [21 ]. Data management and analysis was done using SequenomTyper 4.0 Software. We use the Sequenom MassARRAY RS1000 system to analysis genotype, which was conform to the standard protocol recommended by the manufacturer.

We searched for all SNPs with minor allele frequencies (MAFs) ≥ 0.02 within the region of the MDM4 gene in the 1000 Genomes Project Chinese Han Beijing population database. MAF ≥ 0.02 and tagging r² ≥ 0.8 were used as a screening standard in the selection of tag SNP, which generated 24 tag SNPs for our study. As a result, these 24 tag SNPs (rs3014610, rs2169137, rs117139931, rs137991330, rs4252707, rs190876924, rs12024619, rs72644182, rs117137314, rs76605997, rs76432362, rs116854458, rs12138846, rs61421373, rs191840558, rs116907825, rs115517182, rs12567161, rs150337092, rs80242302, rs3789044, rs3789043, rs884108 and rs61817485) were included in further analyses. All our selected SNPs had P values greater than 0.05 by the HWE test. Commercial kits were used to extract genomic DNA from peripheral blood leukocytes (Genomic DNA kit, Axygen Scientific Inc., CA, USA). Genotyping was conducted for 24 selected SNPs by using the platform of Sequenom Mass ARRAY RS1000 system (Sequenom, San Diego, CA, USA). Typer Analyzer software (Sequenom, San Diego, California, USA) was used to process signal results to ultimately generate genotype data^{20 (link)}. Case and control statuses were blinded during all genotyping processes for quality control. Five percent of the random samples were repeated, and the results were 100% concordant.