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abc2207154

新虫 (初入文坛)

[求助] 藻类单细胞PCR

谁会藻类的单细胞PCR,我吸取藻类不同细胞数做P,扩增不出来啊。
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blackrose8089

铁杆木虫 (职业作家)

【答案】应助回帖

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小丹木木: 金币+2, 鼓励交流 2012-11-08 13:43:14
衣藻的一个方法,供参考:


To quickly and easily screen one or several strains of Chlamydomonas the following protocol works amazingly well:


1) resuspend a pellet of approximately 5-10 uL in size in 50 uL of 10mM NaEDTA by vortexing.
2) incubate at 100C for 5 minutes (thermocycler works well).

3) vortex to mix.

4) centifuge at 12000g for 1 minute.

5) use 1.5 uL in a 25 uL PCR reaction.
jiayoubashaonian
4楼2012-11-08 10:42:05
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cicelyzh

铁杆木虫 (职业作家)

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藻本身没有细菌好破,所以单藻落PCR有时候会不成功。一般都是从平板上刮若干细胞再用酚氯仿抽提出总DNA后做PCR比较保险。
2楼2012-11-07 08:22:32
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abc2207154

新虫 (初入文坛)

引用回帖:
2楼: Originally posted by cicelyzh at 2012-11-07 08:22:32
藻本身没有细菌好破,所以单藻落PCR有时候会不成功。一般都是从平板上刮若干细胞再用酚氯仿抽提出总DNA后做PCR比较保险。

我现在做的是单细胞PCR啊
3楼2012-11-08 09:41:28
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Misszhu

金虫 (著名写手)

引用回帖:
4楼: Originally posted by blackrose8089 at 2012-11-08 10:42:05
衣藻的一个方法,供参考:


To quickly and easily screen one or several strains of Chlamydomonas the following protocol works amazingly well:


1) resuspend a pellet of approximately 5-10 uL in  ...

大神,请问为什么用NaEDTA溶解藻细胞呢?
5楼2020-04-26 21:47:59
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