Simultaneous DNA'fingerprinting', diagnosis of sex and single-gene defect status from single cells

Sex and cystic fibrosis status have been previously diagnosedseparately at the single cell level. We have developed a sensitive,reliable, accurate and rapid (within 5–6 h) system forthe simultaneous diagnosis of sex, cystic fibrosis and a DNA‘fingerprint’ within a single reaction from a varietyof single cells. As contamination cannot be totally excluded,particularly at the single cell level, DNA ‘fingerprinting’can be used to assess the risk of contamination. High sensitivitywith single cells is combined with very high specificity (estimatedmatching probability of 10^–7–10^–8), allowingthe source of the amplified cell to be identified with a veryhigh degree of probability. Fluorescent primers were multiplexedfor six tetranucleotide microsatellite sequences to determinethe DNA fingerprint; the amelogenin gene was used to diagnosesex, and primers for the CFTR region were used to determinecystic fibrosis (CF) status. Analysis of the fluorescent productwas undertaken using an automated DNA sequencer with Genescansoftware. This technique has many applications such as prenataland preimplantation diagnosis, forensic identification of smallor degraded samples, and detection of contamination sources.DNA fingerprints of single haploid spermatozoa and other cellscan be assessed, so ensuring the detection of both diploid andhaploid contamination during preimplantation diagnosis. cystic fibrosis/DNA fingerprinting/fluorescent PCR/preimplantation diagnosis/sexing