非缺失/重复型Duchenne肌营养不良症患者的致病点突变分析  

Identification of disease-causing point mutations in DMD patients' dystrophin gene without large deletions/duplications

作　　者：申本昌[1] 张成[1] 陈松林[1] 孙筱放 李少英 姚晓黎[1] 王淑辉[1] 卢锡林[1] 

机构地区：[1]中山大学附属第一医院神经科,广州510080 [2]广州市第二人民医院妇科

出　　处：《中华医学遗传学杂志》2006年第4期392-396,共5页Chinese Journal of Medical Genetics

基　　金：国家自然科学基金(30370510);广东省自然科学基金博士启动项目(5300783及04300353);卫生部临床重点项目基金(2001321);高等学校博士学科点专项科研基金(200330558058);中国博士后科学基金(2005037172)

摘　　要：目的检测非缺失/重复突变型Duchenne肌营养不良症(Duchennemusculardystrophy,DMD)的致病点突变。方法对6个家系的6个无关DMD男性患者的DMD基因的79个外显子及5′-、3′-非翻译序列进行PCR扩增,产物通过变性高效液相色谱(denaturinghighperformanceliquidchromatography,DHPLC)技术进行突变筛查。结果6例非缺失/重复突变型Duchenne肌营养不良症患者,检测出了5例患者的致病点突变,即697-698insGT,C616T,G1255T,C4279T和C2302T。第1个点突变引起移码突变,后4个致病点突变引起翻译的提前终止,最终导致Duchenne肌营养不良症。患者3除致病点突变外,在第39内含子还发现1个T5586+61A点突变;患者5还检测出了一个位于第8外显子的错义突变;而没有检出致病点突变的患者6,发现了2个外显子突变及2个内含子序列点突变,即C2168+13T、G5234A、C5280T和5740-13dupG。所有检出的突变有7个点突变未见报道。结论变性高效液相色谱技术结合测序,可用于检测DMD患者的点突变,该方法具有准确、灵敏的特点。Objective To detect the disease-causing point mutations in the dystrophin gene of Duchenne muscular dystrophy （DMD） patients. Methods The approach of denaturing high performance liquid chromatography （DHPLE） coupling with sequencing was used to screen the point mutations of 79 exons and the untranslated regions of dystrophin gene without large deletions/duplications, which was in 6 unrelated DMD probands from 6 DMD families. Resuits Five disease-causing mutations, 697-698insGT, C616T, G1255T, C4279T, and C2302T, were identified in 6 probands. The first one resulted in reading frame change; the others created the new stop codons in downstream sites of mutations, respectively. In addition to the disease-causing point mutations, a point mutation T5586 ＋ 61A in intron 39 was also found at patient 3, and a missense mutation A694T in exon 8 was detected at patient 5. Four point mutations, C2168 ＋ 13T, 5740-13dupG, G5234A and C5280T, were also detected at patient 6 whose causative point mutation was unavailable. Seven point mutations have not been reported previously. Bi-directional PCR amplification of specific alleles （Bi-PASA） method was estabhshed to distinguish the haplotypes of heterozygote or homozygote in a single PCR reaction. Conclusion Via automated DHPI.E screening or detecting the subexonic mutations in dystrophin gene is feasible to clinical laboratories, and also is a superior method in terms of sensitivity and efficiency.

关 键 词：Duchennc肌营养不良症 变性高效液相色谱 双向PCR特异位点扩增 点突变 

分 类 号：R746.2[医药卫生—神经病学与精神病学] R392.11[医药卫生—临床医学]