求高手指导DNA测序实验怎么做具体步骤是怎样的

2025-03-14 23:33:50
推荐回答(1个)
回答1:

历史是 Rober Holley,1965年发表于Science杂志上的文章报道了酵母丙氨酸tRNA序列,77碱基。 吴瑞博士(Dr. Ray Wu)提出引物-延伸的测序策略。1971年首次成功的测定了λ噬菌体的两个粘性末端。 F.Sanger,1977年,发表在Nature和PNAS上的文章描述了双脱氧链终止法。诺贝尔奖获得者。 A.M.Maxam和W.Gilbert,1977年,化学法测序。 双脱氧链终止法测序(又称酶法,Sanger测序方法): 对DNA进行体外合成时,将ddNTP与dNTP按一定的比例混合就可以得到一组大小不同、按照模板序列终止的DNA片段 如果引物是带有标记的,在变性凝胶上做电泳分离,通过放射自显影技术就可以检测单链DNA片段,并进行认读 化学法DNA测序(又称Maxam-Gilbert法) 是一个DNA化学降解的过程,而不是生物合成过程。 化学试剂处理具有末端标记的DNA片段,造成碱基的特异性切割,由此产生一组具有各种不同长度的DNA链的反应混合物。 经凝胶电泳按大小分离和放射自显影后,根据X光片所显现的相应条带,直接读出待测DNA片段的核苷酸顺序。 PS:呵呵,我是说的最主要的小片段测序法,如果要是测比较大的序列,像人类基因组那类,我记得听杨焕明教授说他们是用的鸟枪法,呵呵,要针对具体的实验去选择方法

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