Analysis of a polymorphic microRNA target site in the purinergic receptor P2RX7 gene

The recent discovery of post-transcriptional regulation by microRNAs (miRNAs) drew our attention to SNPs of putative miRNA target sites in candidate genes of depression-related psychiatric disorders. The P2RX7 (purinergic receptor P2X, ligand-gated ion channel, 7) gene has been suggested as a candidate for major depressive and bipolar disorder, because of repeated associations with the rs2230912 (G1n460Arg) polymorphism. As this polymorphism is located at the end of the coding region, we considered a possible linkage with SNP(s) in putative miRNA target sites of the 3' untranslated region. Based on our in silico search, the rs1653625 fulfilled this criterion. This SNP, however, is surrounded with polycytosine and polyadenine tracts, which hindered its analysis until now. In this study, we describe a readily applicable genotyping method for rs1653625 by applying a primer that introduces mismatched nucleotides to create a restriction enzyme cleavage site. The resulting allele-specific products with 19 base pair difference were separated by both traditional horizontal agarose gel electrophoresis and multicapillary gel electrophoresis. The developed genotyping method was applied in our depression-related association study.