应用限制性酶谱分析法快速检出Wilson病基因突变热点  

Rapid detection of the hot point mutations of Chinese Wilson disease's gene with PCR-based restriction fragment length polymorphism

作　　者：王柠[1] 吴志英[1] 林珉婷[1] 慕容慎行[1] 

机构地区：[1]福建医科大学附属第一医院神经内科,福州350005

出　　处：《中华神经科杂志》1999年第5期281-283,共3页Chinese Journal of Neurology

基　　金：卫生部优秀青年人才专项基金!97082;福建省卫生厅基金!96A032

摘　　要：目的建立Wilson病（WD）基因突变热点的快速检出方法，并探讨其在WD临床可疑患者诊断中的价值。方法PCR扩增我国WD基因的突变热区──第8号和12号外显子，分别以限制性内切酶MspI和TaiI消化扩增产物，2％琼脂糖凝胶电泳分离，得出相应限制性酶切图谱并进行分析。56例非同源家系的WD患者及60例正常对照进行了该项检测。其中44例同时进行这两个外显子的测序检测。结果37．5％（21／56）的WD患者在第8号外显子检测到Arg778Leu／Gln点突变，其中12例为纯合点突变，9例为杂合点突变，染色体突变频率为29．5％（33／112）。16．1％（9／56）的患者在第12号外显子检测到Thr935Met点突变，均为杂合点突变，染色体突变频率为8．0％（9／112）。结果与测序相符。结论采用限制性酶谱分析法可准确检出WD基因最常见的两个突变点，有助于对临床可疑患者进行诊断。并具有简便快速、结果清晰可靠、不需用同位素等优点，易于推广应用。Objective To establish a rapid method to detect the hot point mutations of Wilson disease(WD) gene and to investigate its clinical application value. Method Arg778Leu/Gln of exon 8 and Thr935Met ofexon 12 were hot point mutations of Chinese WD gene. They were amplified by polymerase chain reaction (PCR),analyzed by digestion with restriction enzymes MspI and Tail, respectively, and followed by 2% gel electrophoresisof the cleavage products . 56 unrelated WD patients and 60 unrelated normal controls were screened with thismethod. Results Arg778Leu/Gln of exon 8 was detected in 37. 5% (21/56)patients. Among them, 12 patientswere homozygous and another 9 patients were heterozygous for this mutation. The frequency of the mutation was29.5% (33/112). Thr935Met of exon 12 was detected in 16.1% (9/56) patients and 9 cases were allheterozygous for this mutation. The frequency of the mutation was 8. 0% (9/112). The result of this method was thesame as that of the DNA sequencing. Conclusion A direct diagnostic system based on the method described herewould, therefore, be expected to identify the two most common mutations of Chinase WD gene and help to diagnoseWD cases suspected on clinical grounds. The advantage of the method would be that no radioactive PCR andelectrophoresis were needed, and that the results were clear, and unambiguous. With this method, it would bepossible to diagnose the WD patients accurately, conveniently and rapidly.

关 键 词：肝豆状核变性 基因突变 限制酶图谱法 

分 类 号：R742.402[医药卫生—神经病学与精神病学]