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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