毛细管电泳法检测肝细胞癌微卫星基因表型的影响因素

目的：分析影响肝细胞癌微卫星—自动毛细管电泳检测结果的技术因素。方法：应用MegaBACE500毛细管电泳测序仪对一组高度多态性微卫星PCR扩增产物进行电泳，应用Genetic Profiler软件进行基因表型分型，比较PCR扩增产物在不同处理条件下对基因分型的影响。结果：PCR反应体系残留混合物(dNTP、引物和盐离子的浓度等)对基因分型的结果有明显的影响，当PCR体系中益离子和DNA的浓度比＞10000：1时可使等位基因片段分析系统评分出现偏差而导致结论错误，基因组DNA—PCR样品浓度在50ng/μl时较为合适。结论：毛细管电泳测序体系中PCR反应体系残留混合物的浓度是影响基因分型结果的一个重要因素。

Influencing factors of capillary electrophoresis system in automatic microsatellite genotyping of hepatocellular carcinoma

Objective: To study technique influences on microsatellite-automatic capillary array electrophoresis in hepa-tocellular carcinoma. Methods: A panel of hyper-polymorphic microsatellites were detected using MegaBACE-500 capillary array electrophoresis (CEA) instrument, and were analyzed with MegaBACE Genetic Profiler software. Influences of PCR reaction residues at different concentrations on genotyping were investigated. Results: The concentration of PCR reaction residues (including dNTP, primers and salt ion) had obvious effect on scoring errors in allele size. When the ratio of salt ion: DNA in PCR system was >10 000 : 1, the scoring might result in misleading conclusions. The concentration of genomic DNA-PCR reaction products in 50 ng/fd was suitable for CEA analysis. Conclusion: The concentration of PCR reaction residues in CEA system is one of the key factors altering genotyping results.