利用单线虫多重PCR技术快速制备DNA分子量标准

A Method for Rapid Preparation of DNA Molecular Weight Standard by Single C. elegans Multiplex PCR

基于单只线虫6对染色体及多重PCR技术建立一种特异性扩增特定长度核酸片段的方法,进而探索其在常规核酸分子量标准制备中的应用。利用多重PCR引物设计及特异性评估软件以秀丽线虫6对染色体为模板,分别设计扩增不同长度核酸片段的特异性引物对,并直接对单只秀丽线虫进行多重PCR扩增,采用20g/L琼脂糖凝胶电泳进行分离鉴定并采集图像,通过系统研究优化获得特异性高的1～6重PCR扩增产物,产物大小经鉴定与预期目的条带一致,6重PCR扩增产物条带与常规分子量标准DL 2 000相应条带完全吻合,说明采用该方法能够有效制备特定片段大小的分子量标准。

In order to establish a single C.elegans multiplex PCR method to amplify specific DNA fragment and further exploring its application in preparing of nucleic acid molecular weight standards,six pair primers were designed according to the six chromosome DNA sequences of C.elegans and the MP primer and MFE primer programs were used to evaluate their specificities.Subsequently,a single nematode was used as the template to run multiplex PCR,followed with further identification by 20 g/L agarose gel electrophoresis.Different multiplex PCR products were obtained as expected.The six-primer multiplex PCR products are identical to the widely used nucleic acid molecular weight standard DL 2 000.Our method can be used to prepare different nucleic acid molecular weight standards.In summary,a fast and specific multiplex PCR method was developed and different nucleic acid molecular weight standards could be obtained by this method.This method provides a reference for further using of multiplex PCR.