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目的 探讨常染色体显性遗传性多囊肾病 (autosomal dominant polycystic kidney disease,ADPKD)的症状前基因诊断方法。方法 PCR扩增 PKD1基因两侧和基因内 SM6、16 AC2 .5、HBAP、KG8共 4个微卫星遗传标记 ,并采用中性聚丙烯酰胺电泳 (polyacrylamide gel electrophoresis,PAGE)凝胶、常规变性 PAGE凝胶、及含 32 %甲酰胺、5 .6 mol/ L 尿素的 PAGE凝胶进行电泳检测。结果 被检测家系疾病与 PKD1连锁。在 SM6位点上 ,中性胶检测 4 和 1 均为纯合子 ,在变性胶检测中则是杂合子。在常规变性 PAGE凝胶电泳中 ,SM6位点扩增产物有很多杂带干扰结果分析 ,而在含 32 %甲酰胺、5 .6 mol/ L尿素电泳检测中无杂带干扰 ,主带很清楚。结论 联合应用多个微卫星标记对 ADPKD进行连锁分析能有效地进行早期诊断。与中性胶相比 ,采用变性胶对微卫星标记 PCR扩增产物进行电泳检测 ,所获得的信息含量高 ;而采用含 32 %甲酰胺、5 .6 mol/ L 尿素的聚丙烯酰胺凝胶更能有效地消除杂带 ,优于常规变性胶。
Objective To investigate the method of presymptomatic gene diagnosis of autosomal dominant polycystic kidney disease (ADPKD). Methods Four microsatellite markers, SM6, 16 AC2. 5, HBAP and KG8, on both sides of the PKD1 gene and in the gene were amplified by PCR. Polyacrylamide gel electrophoresis (PAGE) gels and conventional denaturing PAGE Gel and PAGE gel containing 32% formamide and 5.6 mol / L urea for electrophoresis. The results were tested pedigree disease and PKD1 chain. At the SM6 locus, Neutral Glue Assay 4 and 1 are both homozygous and heterozygous in denaturing Gum Assay. In the conventional denaturing PAGE gel electrophoresis, SM6 locus amplified products have a lot of grizzly interference results analysis, and in 32% formamide, 5.6 mol / L urea electrophoresis test without interference, the main band is clear . Conclusion The combination of multiple microsatellite markers for linkage analysis of ADPKD can effectively diagnose early. Compared with the neutral gel, the denatured gel microsatellite markers PCR amplification products electrophoresis detection, obtained high content of information; and with 32% formamide, 5.6 mol / L urea polyacrylamide gel More effective to eliminate miscellaneous goods, better than conventional denatured plastic.