(by R.Jayakumar from roswellpark.org)
Tue Dec 22 15:45:58 EST 2009
You mean you want to linearise the construct by cutting it with a RE outside the expression cassette before transformation??? Is that what you mean. I don't see the purpose of cutting a vector after ligating vector and fragment to clone a PCR fragment efficiently. It does not make much sense.
From: methods-bounces from oat.bio.indiana.edu [mailto:methods-bounces from oat.bio.indiana.edu] On Behalf Of Nasser Mahna
Sent: Tuesday, December 22, 2009 12:23 PM
To: methods from magpie.bio.indiana.edu
Subject: Cloning trick
I would to ask if there is anybody mastered in working with restriction and digestion.
I want to clone a pcr fragment inside a binary vector. To do it efficiently, I want to use a restriction enzyme after ligating vector and fragment, which cuts the original uncut vector before transformation.
what method do you offer?
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