A flexible SNP genotyping system to study allelic determinants of brain function

We developed a system for SNP-genotyping consisting of two ‘‘microchips’’, one investigating 40 SNPs in genes involved in the hepatic metabolism of antidepressant drugs, the other 100 SNPs in genes involved in brain response to antidepressants. Candidate genes were chosen by literature screening. SNPs were selected on HapMap data, using the pairwise algorithm combinedwith the ‘‘BestN’’ method; a few SNPs, reported as associated with treatment outcome, were added to the set. The main advantages of our experimental design are: a) simultaneous analysis of several genes (22) involved in distinct aspects of antidepressant response; b) a comprehensive search for common variability in candidate genes by HapMap ‘‘tag-SNPs’’. DNA samples are genotyped by ‘‘padlock probes’’, which specifically recognise the two variants of each SNP; padlock probes are amplified with universal primers, and contain unique tags for hybridisation to known positions on ad hoc designed micro-arrays, where the two alleles are distinguished by distinct fluorophores. Since reactions carried out on multiplex and signal intensities on microarrays are homogeneous, as they are not affected bySNPflanking sequences, tens of SNPsfrom tens of individuals can be analysed simultaneously on a single slide, with high accuracy and reproducibility. Considering that 100 SNPs from manygenes related to neural transmission, plasticity and neurogenesis are included in the secondmicrochip and that each of them can easily be substituted withothers if needed, our system represents a powerful tool for investigating genetic influence on normal and pathological brain function.