论文部分内容阅读
目的探讨和优化基于微流控技术的DNA片段分离技术用于伤寒沙门菌多位数目可变串联重复序列分析(MLVA)的方法。方法制备与分析片段长度相似的内标来提高实验结果分析的重复性;用Agilent 2100分析仪和序列测定技术分析伤寒沙门菌10个数目可变串联重复序列(VNTR)位点的PCR产物,比较2种方法的一致性,优化基于微流控的DNA片段分析技术。结果新内标能够提高方法的重复性;使用Agilent 2100分析仪电泳以及新内标校正片段后,具备相同重复单元拷贝数的VNTR扩增产物具有相同的电泳位置,通过序列测定证实了这些分析结果。结论优化的基于微流控技术的DNA片段分离方法能够提高短重复序列MLVA分析的准确性,可应用于伤寒沙门菌的MLVA分析。
Objective To explore and optimize the method of DNA fragmentation based on microfluidic technology for the analysis of multiple variable number of Salmonella typhi by MLVA. Methods An internal standard with similar fragment length was prepared to improve the repeatability of the experimental results. PCR products of 10 variable tandem repeat (VNTR) sites of Salmonella typhi were analyzed by Agilent 2100 analyzer and sequence analysis. The consistency of the two methods optimizes microfluidic-based DNA fragment analysis techniques. Results The new internal standard improved reproducibility of the method; VNTR amplification products with the same copy number of repetitive units had the same electrophoresis position after electrophoresis with the Agilent 2100 analyzer and the new internal standard correction fragment, which were confirmed by sequence analysis . Conclusion The optimized DNA fragmentation method based on microfluidic technology can improve the accuracy of MLVA analysis of short repeat sequences and can be applied to MLVA analysis of Salmonella typhi.