Duchenne型肌营养不良症的产前基因诊断研究

A study of the prenalal gene diagnosis in Duchenne muscular dystrophy

应用Ｄｕｃｈｅｎｎｅ型肌营养不良症（Ｄｕｃｈｅｎｎｅ ｍｕｓｃｕｌａｒ ｄｙｓｔｒｏｐｙ，ＤＭＤ）基因的ｃＤＮＡｓ作为探针，以限制性片段长度多态（ＲＦＬＰ）分析为策略，采用Ｓｏｕｔｈｅｒｎ分子杂交方法，成功地对１例可疑ＤＭＤ的男性胎儿及１例ＤＭＤ患儿进行基因诊断。结果显示该胎儿ＤＭＤ基因正常，而患儿存在ＤＭＤ基因缺失（缺失２．１５ｋｂ）。在基因分析前，应用聚合酶链式反应（ｐｏｌｙｍｅｒａｓｅ ｃｈａｉｎ ｒｅａｃｔｉｏｎ，ＰＣＲ）技术鉴定胎儿性别为男性。胎儿出生后检查结果与与产前基因诊断相吻合。为了获得高灵敏度探针，本文采用地高辛配基标记ＤＮＡ探针的方法，通过酶联免疫法，使分子杂交的ＤＮＡ检测带出现颜色反应。实验结果表明，此方法适用于基因组单拷贝ＤＮＡ顺序的检测，具有快速、安全等优越性，可以替代同位素进行推广、应用。

We used cDNA8 probe cloned from DMD cDNA(14 Kb)and Southern molecular hybridization technique and adopted the analysis strategy of restriction fragment length polymorphisms(RF-LPs).We successfully made gene diagnosis on one male fetus suspected with DMD and a DMD boy, and discovered a DMD gene deletion(2.15kb)on the boy of this family pedigree. Meanwhile,a normal gene was found on the fetus. Before the gene was analysed, we had the fetus sex determined using polymerase chain reaction(PCR)technique and male fetus was recognized. The result of a postnatal examination on this fetus coincided with our prenatal gene diagnosis. In order to obtain the DNA labeling probe with a high sensitivity, we achieved random primed DNA labeling with digoxigenin-dUTP and got a color reaction of the DNA detection band of molecular hybridization through enzyme-link immunoassay.The result showed that our experimental method was suitable for the detection of single copy DNA sequence of the genme, and it had merits of rapidity and safety,and the possi bility of taking place of isotope technique in actual application and propagation.