摘要
目的:制备一种具有琼脂糖凝胶电泳定量功能的DNA分子量标准。方法:以pMD18-TSimple载体为基础骨架构建了长度为4.7kb的质粒,应用定点突变的方法,在载体上分别间隔100bp、200bp、400bp、800bp、1200bp处,加入了HindⅢ限制性内切酶的酶切位点,将该质粒扩增后并应用HindⅢ酶切后,1.5%琼脂糖凝胶电泳鉴定。结果:获得的分子量条带大小依次为100bp、200bp、400bp、800bp、1200bp和2000bp,每次使用4μl可获得质量范围为10ng、20ng、40ng、80ng、120ng和200ng的定量标准品。结论:应用该方法制备标准品,具有制备简单、成本低、定量快速等优点。
Objective：To make a DNA molecular weight marker with estamiting DNA amount on agarose gel electrophorose.Method：A 4.7 kb plasmid was made by inserting a fragment into pMD18-T simple vector.Using site-directed mutagenesis strategy,several HindⅢ retriction enzyme sites were inserted into this vector with interval of 100 bp,200 bp,400 bp,800 bp,1 200 bp and 2000 bp.DNA mass MW marker was prepared by HindⅢ digestion and checked on agarose gels.Result：The band sizes of DNA mass MW marker are 100 bp,200 bp,400 bp,800 bp,1200 bp and 2000 bp,it indicates DNA amount of 10 ng,20 ng,40 ng,80 ng,120 ng and 200 ng respectively,when applying 4μl on agarose gel.Conclusion：The DNA mass MW marker is advantage of made easy,cheap,simple to use.
出处
《生物技术》
CAS
CSCD
北大核心
2010年第3期62-64,共3页
Biotechnology
