SNPs were selected using the dbSNP database ( http://www.ncbi.nlm.nih.gov/projects/SNP/ ) and were chosen based on functional significance. Minor allele frequency according to 1000Genomes database is 0.41 for rs4880, 0.48 for rs7943316, 0.09 for rs1800629, and 0.14 for rs1800795.
All SNPs were analyzed with the Polymerase Chain Reaction-Restriction Fragment Length Polymorphism technique (PCR-RFLP). Samples were amplified in 15 μL reactions containing 7.5 uL of GoTaq® G2 Hot Start Green Master Mix ( Promega Corporation, St. Madson - USA ), 1 μL of each primer (10 μM), 1 μL of DNA, and nuclease-free water. Primer sets as well as the enzymatic digestion conditions were the same as previously described.16 (link) , 18 (link) , 24 (link) Genotypes were analyzed through vertical electrophoresis in 6% polyacrylamide gels, followed by coloring with silver nitrate. The genotypes were identified by their band pattern according to literature.21 (link) , 25 (link) , 31 (link)SOD2 restriction fragments: CT (246, 157, 89 bp) and TT (157, 89 bp); CAT restriction fragments: TT (249 bp), AA (175, 74 bp), and AT (249, 175, 74 bp); TNF-α restriction fragments: GA (107, 87 bp), GG (107bp), and AA (87 bp); and IL-6 restriction fragments: CC (117bp), GG (139bp), and GC (139, 117bp).
All SNPs were analyzed with the Polymerase Chain Reaction-Restriction Fragment Length Polymorphism technique (PCR-RFLP). Samples were amplified in 15 μL reactions containing 7.5 uL of GoTaq® G2 Hot Start Green Master Mix ( Promega Corporation, St. Madson - USA ), 1 μL of each primer (10 μM), 1 μL of DNA, and nuclease-free water. Primer sets as well as the enzymatic digestion conditions were the same as previously described.16 (link) , 18 (link) , 24 (link) Genotypes were analyzed through vertical electrophoresis in 6% polyacrylamide gels, followed by coloring with silver nitrate. The genotypes were identified by their band pattern according to literature.21 (link) , 25 (link) , 31 (link)SOD2 restriction fragments: CT (246, 157, 89 bp) and TT (157, 89 bp); CAT restriction fragments: TT (249 bp), AA (175, 74 bp), and AT (249, 175, 74 bp); TNF-α restriction fragments: GA (107, 87 bp), GG (107bp), and AA (87 bp); and IL-6 restriction fragments: CC (117bp), GG (139bp), and GC (139, 117bp).
Free full text: Click here
