Fluorescence in situ hybridization (FISH) validation of the GRM5 deletion was performed on metaphase spreads prepared from the participant-derived lymphoblastoid cell lines using standard methodology. Chromosomes were visualized by counter staining with 4,6-diamidino-2-phenylindole. FISH was performed using fosmid W12-2219g4 and BAC RP11-697e14. FISH analysis was carried out as previously described.28 Bacterial artificial chromosomes and fosmids were isolated using the UltraClean plasmid kit (MoBio Labs Inc., Carlsbad, CA, USA) and probes were labeled with Spectrum Red or Green (Abbott Molecular Inc., IL, USA) by nick translation. FISH images were captured using MacProbe software (Applied Imaging, San Jose, CA, USA).
