试剂盒配合琼脂糖EB染色法和PAGE银染法分析线粒体DNA A1555G突变的比较研究

Mitochondrial DNA A1555G Mutation Analyzed by Testing Kit with PAGE Silver Staining and Ethidium Bromide Staining Methods

李琦;戴朴;黄德亮;金政策;康东洋;张昕;

1:解放军总医院耳鼻咽喉头颈外科解放军总医院耳鼻咽喉科研究所解放军总医院聋病分子诊断中心

2:解放军总医院耳鼻咽喉头颈外科解放军总医院耳鼻咽喉科研究所解放军总医院聋病分子诊断中心

3:解放军总医院耳鼻咽喉头颈外科解放军总医院耳鼻咽喉科研究所解放军总医院聋病分子诊断中心

4:山东省威海市威海澳麦尔基因科技有限公司

摘要
目的通过比较试剂盒配合琼脂糖EB染色法和PAGE银染法分析线粒体DNAA1555G突变的不同特点,探索应用标准试剂盒和PAGE银染方法检测线粒体DNAA1555G突变的快速筛查和诊断方法。方法采用线粒体DNAA1555G突变诊断试剂盒配合PAGE胶电泳银染法检测线粒体DNA1555位点正常者229例,A>G突变阳性者17例,验证此试剂盒配合PAGE银染法检测的有效性和准确性。结果线粒体DNAA1555G突变诊断试剂盒配合PAGE胶电泳银染法检测结果与酶切EB染色和测序结果完全吻合。结论线粒体DNAA1555G突变诊断试剂盒配合PAGE胶电泳银染法和试剂盒配合EB检测法较前者更为灵敏和清晰,在分析线粒体DNAA1555G突变方面具有简单、价廉、结果直观的特点,适合在中国用于进行此突变的大规模筛查或预防性检查。
关键词
耳聋;线粒体DNA;突变;聚丙烯酰胺凝胶电泳;银染;溴化乙锭染色
基金项目(Foundation):
作者
李琦;戴朴;黄德亮;金政策;康东洋;张昕;
参考文献

1Prezant T,Agapian G,Bohl man M,et al.Mitochondrialribo-somal RNA mutation associated with both antibiotic-induced and non-syndromic deafness[J].Nat Genet,1993,4:289.

2Li Z,Li R,Liao Z,et al.Mutational analysis of the mitochon-drial12S rRNA gene in Chinese pediatric subjects with amin-oglycoside-induced and non-syndromic hearing loss[J].Hum Genet,2005,117:9.

3Bacino C,Prezant T,Bu X,et al.Susceptibility mutations in the mitochondrial small ribosomal RNAgenein aminoglycoside induced deafness[J].Pharmacogenetics,1995,5:165.

4Tessa A,Giannotti A,Tieri L,et al.Maternally inherited deaf-ness associated with a T1095C mutationinthe mtDNA[J].Eu-ro J Hum Genet,2001,9:147.

5Hutchin T,Parker M,Young I,et al.A novel mutation in the mitochondrial tRNA(Ser(UCN))genein a family with non-syndromic sensorineural hearing i mpairment[J].J Med Genet,2000,37:692.

6Sue C,Tanji K,Hadjigeorgiou G,et al.Maternally inherited hearing loss in a large kindred with a novel T7511C mutation in the mitochondrial DNAtRNA(Ser(UCN))gene[J].Neu-rology,1999,52:1905.

7Pandya A,Xia X,Erdenetungalag R,et al.Heterogenous point mutations in the mitochondrial tRNASer(UCN)precursor co-existing withthe A1555G mutationin deaf students from Mon-golia[J].AmJ Hum Genet,1999,65:1803.

8Guan M,Enriquez J,Fischel-Ghodsian N,et al.The deaf-ness-associated mitochondrial DNA mutation at position7445,which affects tRNASer(UCN)precursor processing,has long-range effects on NADHdehydrogenase subunit ND6gene expression[J].Molecu Cellu Bio,1998,18:5868.

9Smith R,Bakejr J,White K.Sensorineural hearing loss in children[J].Lancet,2005,365:879.

10del Castillo F,Rodriguez-Ballesteros M,Martin Y,et al.Heteroplasmy for the1555A>G mutationin the mitochon-drial12S rRNAgene in six Spanish families with non-syn-dromic hearing loss[J].J Med Genet,2003,40:632.

11el-Schahawi M,Lopez de Munain A,Sarrazin A,et al.Twolarge Spanish pedigrees with nonsyndromic sensorineural deafness and the mtDNA mutation at nt1555in the12S rRNAgene:evidence of heteroplasmy[J].Neurology,1997,48:453.

12Fischel-Ghodsian N.Mitochondrial deafness[J].Ear Hear,2003,24:303.

13戴朴,杨伟炎,韩东一,等.Prev-DAF试剂盒分析线粒体基因1555A-G突变[J].中华耳科学杂志,2004,2:37.

14Li X,Greinwald J,Yang L,et al.Molecular analysis of mi-tochondrial12SrRNAandtRNASer(UCN)genesin pediatric subjects with nonsyndromic hearing loss[J].J Med Genet,2004,41:615.

15Fischel-Ghodsian N,Prezant T,Chaltraw W,et al.Mito-chondrial gene mutationis a significant predisposing factor in aminoglycoside ototoxicity[J].AmJ Otolaryngol,1997,18:173.

16Usami S,Abe S,Akita J,et al.Prevalence of mitochondrial gene mutations among hearing i mpaired patients[J].J Med Genet,2000,37:38.

17戴朴,刘新,于飞,等.全国18个聋校学生非综合征性聋分子病因学研究(Ⅰ)[J].中华耳科学杂志,2006,4:1.

18张劲,李琦,戴朴,等.乌鲁木齐市特教学校重度感音神经性聋GJB2和线粒体基因常见突变调查[J].中华耳科学杂志,2007,5:60.