基于微流控的核酸片段分离技术用于伤寒沙门菌MLVA分型的研究

王洪霞; 崔志刚; 熊礼凤; 章丽娟; 阚飙

doi:10.3784/j.issn.1003-9961.2009.03.021

基于微流控的核酸片段分离技术用于伤寒沙门菌MLVA分型的研究

Study on multiple locus VNTRs analysis of <I>Salmonella</I> Typhi by nucleic acid separation technology based on microfluidics

摘要

摘要: 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。

Abstract: ObjectiveTo probe into and optimize the technology of multiple locus VNTRs analysis of ISalmonella/I Typhi by microfluidics-based nucleic acid separation technology. MethodsTo improve reproducibility of experiment by artificial internal markers which resemble DNA fragment analyzed. To analyze amplification products of 10 ISalmonella/I Typhi VNTR locus by Agilent 2100 analyzer and sequence. And then to compare two methods and optimize microfluidics-based nucleic acid separation technology. ResultsThe new internal markers could increase reproducibility of experiment. After electrophoresis by Agilent 2100 bioanalyzer and revise by novel internal marker, the location of ISalmonella/I Typhi strains is same on the electropherogram, the VNTR copy numbers of corresponding strains are same. The result was proved by sequence analysis. ConclusionThe optimized DNA separation technology based on microfluidics can use for Multiple locus VNTRs analysis of ISalmonella/I Typhi.

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