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发信人: neverthink (nevernetbug), 信区: Biology
标 题: Re: 请教一个ligation的问题
发信站: BBS 未名空间站 (Wed Dec 7 15:59:00 2005)
well, it depends. How big are your fragments? Are u going to use bluescript or
what?
I have done this easily with three pieces before. Actually, one of them was
even blunt ended. But my fragments were not big. both of them less than 300bp
. I recall a guy mentioned he could do three or four big fragment ligations in
one website. Search on line, see whether you can find it. Surely there are
tricks for cloning or magic hands:) as some guy said earlier. Good luck.
BTW, sometimes, shufflings between vectors to bypass this kind of problem
might be even quicker if you are not very family with cloning. Also there are
tons of vectors avalaible to choose too. Ur boss is big and rich, so...
【 在 windysea (甜心小猪~祝福我们的宝贝小雨瞳) 的大作中提到: 】
: 用三个不同的restriction enzymes,
: 把两个pcr fragments接到vector上，
: 如果一次性地做ligation (mix 3 fragments in 1 reactio8225 1] PaiMei ALLOC: [1685 17734]
[08/16 20:25:23 22663 1] guest ENTER [email protected] (ALLOC 4869) [www]
[08/16 20:25:23 7603 1] guest ENTER [email protected] (ALLOC 10176) [www]
[08/16 20:25:23 24068 1] guest ENTER [email protected] (ALLOC 18146) [www]
[08/16 20:25:24 4430 1] guest ENTER [email protected] (ALLOC 3712) [www]
[08/16 20:25:24 7566 1] guest ENTER [email protected] (ALLOC 4384) [www]
[08/16 20:25:24 32174 1] guest ENTER [email protected] (ALLOC 576) [www]
[08/16 20:25:25 27360 1] guest ENTER [email protected] (ALLOC 8737) [www]
[08/16 20:25:25 4433 1] guest ENTER [email protected] (ALLOC 5281) [www]
[08/16 20:25:25 2416 1] guest ENTER [email protected] (ALLOC 2595) [www]
[08/16 20:25:26 6469 1] guest ENTER [email protected] (ALLOC 17345) [www]
[08/16 20:25:26 6475 1] guest ENTER [email protected] (ALLOC 576) [www]
[08/16 20:25:26 24602 1] guest ENTER [email protected] (ALLOC 28576) [www]
[08/16 20:25:26 4767 1] guest ENTER [email protected] (ALLOC 576) [www]
[08/16 20:25:27 8210 1] usdgood ENTER @68.181.225.168
[08/16 2n), efficiency怎样？
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※ 来源:·BBS 未名空间站 http://mitbbs.com·[FROM: 71.127.]
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