Establishment of the method for prenatal diagnosis of genetic disease using DNA probes and chorionic villus sampling

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Carrier detection and prenatal diagnosis of Duchenne muscular dystrophy(DMD) were performed combined with the use of three intragenic genomic probes and chorionic villus sampling in early pregnancy. Total of 13 families with at least one DMD were analyzed, 5 were for carrier detection, the rest 8 for prenatal diagnosis. DNA was extracted from peripheral white blood cells for carrier testing (90 individuals). For prenatal detection, it was extracted from chorionic villi obtained by chorionic villus sampling at 9 menstrual weeks(8 fetuses). DNA was digested with appropriate restriction enzyme followed by overnight electrophoresis into 1% agarose gels. DNA was transferred from the gel to nylon membrane according to a protocol of alkaline transfer method. The PERT 87 probes were labeled by nick translation to a specific activity of 0.7 to 1.3 x 10^9cpm/mug. The membranes were hybridized 15 hours at 41 ﾟC after 4 hours prehybridization. After washing the semi-dried membranes were exposed to X-ray films to make autoradiograms for restriction fragment length polymorphisms analysis. In instances of prenatal diagnosis fetal sex was determined by a rapid screening test with a Y chromosome-specific repeat sequence in band Yq12. Of the 5 at-risk females in 5 families for scarier detection 2 were diagnosed as carriers, 3 as non-carriers. Out of 8 fetuses from 8 families for prenatal diagnosis 4 were males and 4 were females. All of 4 male fetuses were determined to be unaffected. Of 4 female fetuses, 3 were diagnosed as non- carrier, carrier status of the remaining one was not derided because her mother was not informative for all testings. The method for prenatal diagnosis of DMD in early pregnancy was established. This method is applicable to other genetic diseases.