NOVEL AND IMPROVED MOLECULAR MARKERS AND DIAGNOSTIC ASSAYS FOR PREIMPLANTATION GENETIC DIAGNOSIS OF a- AND ß-THALASSEMIA AND FRAGILE X SYNDROME

Abstract

PREIMPLANTATION GENETIC DIAGNOSIS (PGD) OF ALPHA- AND BETA-THALASSEMIA AND FRAGILE X SYNDROME (FXS) IS USUALLY PERFORMED BY DIRECT MUTATION DETECTION OR FLANKING MARKERS. CURRENTLY, LIMITED NUMBERS OF LINKED MARKERS LIMIT THE UTILITY OF LINKAGE-BASED PGD FOR THESE DISEASES. WE PERFORMED IN SILICO MINING TO IDENTIFY POLYMORPHIC MARKERS WITHIN 1 MB ON EITHER END OF THESE GENE LOCI, RESPECTIVELY. MARKERS WITH GOOD HETEROZYGOSITY AND POLYMORPHISM INFORMATION CONTENT WERE CO-AMPLIFIED IN A SINGLE PANEL FOR EACH DISEASE. THE 9-PLEX HBA STR SET AND 15-PLEX HBB STR SET CAN BE FURTHER MULTIPLEXED WITH THE GENE EXONIC FRAGMENTS FOR SIMULTANEOUS DELETION/POINT MUTATION DETECTION WITH LINKAGE ANALYSIS. THE 13-PLEX FMR1 STR SET CAN BE ANALYZED IN PARALLEL WITH FMR1 CGG REPEAT EXPANSION MUTATION DETECTION, USING ALIQUOTS OF WHOLE GENOME AMPLIFIED PRODUCT. ALL THREE SINGLE-TUBE MULTIPLEX-PCR STR ASSAYS HAVE BEEN OPTIMIZED FOR USE ON GENOMIC DNA, SINGLE CELLS, AS WELL AS WGA PRODUCTS OF SINGLE CELLS.