发信人: squidsss (湖心草深長), 信区: Biology
标 题: Re: 求助: Southern Hybridization with olig
发信站: Unknown Space - 未名空间 (Fri Aug 6 23:17:35 2004) WWW-POST

It could be. I do think this method is too sensitive, so it has many critical
parameters (p8 of protocol). If you have one mismatch parameter, the result
screw-up.
I sugest bigfatliar do the dot blot control test first. You just have to drop
several dots of genomic DNA on the membrane, using different concentration.
For example, 1ul of DNA in 10ug/ul, 1ul of DNA in 5ug/ul.... You can have 5
drops in a roll, followed by cross link, hybridization, etc. If this control
membrane worked, then your probe is ok. The problems could be the others.
And another suggestion is change to other labeling methods, either radioactive
or DIG. But I am not sure whether they will work for such short probes.
The Southern is trickiest exp I have ever done. Once you success, you got
everything. But the way to the success is bumpiness.
Good luck to bigfatliar...

【 在 moso (野鬼) 的大作中提到: 】
: I got. Thanks.
: Well, is the ALKPHOS Direct that you used for labeling? I just read the
: instruction and found at the ADDITIONAL INFORMATION that the labeling kit is
: suggested to be used to a probe greater than 50bp in length. Is that mightbe

: a problem?



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※ 修改:·squidsss 於 Aug 6 23:17:35 修改本文·[FROM: 67.39.]
※ 来源:．Unknown Space - 未名空间 mitbbs.com．[FROM: 67.39.]