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Ligation of multi-fragments? - Has anybody tried to ligate more than 02 fragments in a single experiment? (May/20/2006 )

Hi all,

It has been a while I have not visited this forum due to moving! I have read a couple of pages of this forum and found that many questions are directed to the ligation problems.

I am just wondering if anyone out here has tried to ligate 3 or 4 DNA fragments together in a single tube? and if you have tried, what results did you get? is there any special attention should be made to increase the ligation efficiency?

Nice day!


We do it all the time with three fragments. Standard protocols work fine.


Thanks for your reply.

I tried once after reading your reply but few colonies I got do not contain my inserts

I am ligating a 750bp and 200 bp on to pET vector, Do you have any suggestion?