ASSESSMENT OF DIAGNOSTIC QUANTITATIVE FLUORESCENT MULTIPLEX POLYMERASE-CHAIN-REACTION ASSAYS PERFORMED ON SINGLE CELLS

We have refined polymerase chain reaction (PCR) assays for the detecti on of sickle cell anaemia, the delta F 508 deletion causing cystic fib rosis, and the IVS1-110 mutation leading to beta-thalassaemia, allowin g them to be successfully performed upon single cells using fluorescen t primers. We have also assessed the possibility of detecting aneuploi dies of chromosomes 13, 18 and 21 using a quantitative fluorescent pol ymerase chain reaction (QF-PCR) with primers flanking polymorphic shor t tandem repeat (STR) markers. Trisomies were readily diagnosed by the detection of tri-allelic patterns. However some heterozygote normal a nd trisomic diallelic patterns did not produce the expected ratios of amplified PCR products due to preferential DNA sequence amplification. Total allelic drop out (ADO) did not occur with any of the cells test ed. Multiplex QF-PCR assays can be performed on a single cell in under 6 h and simultaneously provide diagnosis of single gene defects, sex determination and an indication of selected chromosome aneuploidy.