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做PCR 时降低污染的小方法

发布: 2016-03-01 748阅读 195评论

1 戴最合适的实验手套：

PCR实验中一个主要的污染来源其实就是实验手套经常接触PCR 管口。即使你觉得自己好像没有碰到，但也给污染物提供了可乘之机。尤其是在实验手套不合手的情况下，小编就见过有人需要带XS的手套，但是XS型号的手套经常存货不多，勉强带S号的，导致实验操作不是很灵巧，容易造成污染。

2在PCR实验前用70% 乙醇擦拭工作台：

我们的皮屑中带有的核酸酶（nuclease）会影响PCR反应的结果。核酸酶例如DNA酶可以切断DNA，影响实验结果。尤其是一个条带都没有的时候，也许就受到了核酸酶的污染。为了避免这样的核酸酶污染以及其他细菌污染，小编建议大家在实验前将台面和实验手套用70%乙醇消毒、擦拭干净。

3加试剂的时候手握PCR管的中下部：

当大家开始做实验的过程中碰到了很多器材，即使是用乙醇擦拭过了，实验手套还是难免不干净。在这种情况下，就要让实验手套尽量远离PCR管口，避免污染物趁机进去。虽然看起来是微不足道的事情，但是往往能直接影响PCR的结果。

4 将试剂分装 （aliquot）：

每次做实验之前，最好将双重去离子水（double deionised water）分装在1ml 或是1.5ml的离心管（eppendorf tube）里。这样每次用完就直接扔掉，保证了去离子水的质量。除此之外，小编建议大家把其他试剂也进行分装，例如缓冲液、聚合酶（polymerase）、dNTPs 和引物（primer）等等。这样可以避免原装瓶中的试剂因为反复使用而被污染的情况。

5仔细关紧PCR管的盖子:

虽然又是看起来很不起眼的一点，但是小编就因为盖子盖得仓促而浪费了宝贵的实验材料和时间。所以希望能够提醒到大家，避免犯简单的错误。如果盖子盖得太仓促，尤其是使用成条PCR 管 （PCR strip）时，很有可能里面的混合物会附着在管壁的上方或是洒出。而且如果盖得太快，没有仔细检查，很有可能没有盖紧，导致在PCR机器里时水分蒸发。这样就会造成高浓度的dNTPs，抑制PCR反应。即使之后的跑胶有结果，也很难保证数据的可靠性和重复性。

6试剂分开放置：

将PCR试剂、产物和其他分子生物学试剂分开放置。例如：一定要用包装袋单独储存PCR管。而且条件允许的话，PCR反应前和反应后的混合物应该放置在冰箱的不同位置。

如果大家有更多、更好的方法，欢迎在平台里留言、交流！

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