Gewtting the concentration of PCR product and cloning

[shifali chatrath]

  Hi!
I want to correct DK's point. If it is proofreading polymerase, 3'A will get depleted off as a result of proof reading. Taq Pol/ Long Pol amplified products can generally be used for TA cloning. Their fact will have that info.

Secondly, to me the suggestion to Point 2 is tedious. 
Gel purified PCR product can be directly quantified and ligation reaction can be set up, depending upon the size of the insert, one needs to clone. 5:1::insert:vector is the optimal molar ratio for cloning insert of 1250bp. Just quantify the insert and go ahead.
All the best.

Shifali



On Thu, 25 Dec 2008 Shri wrote :
>Hi all members,
>I have amplified PCR product of 1250 bp from Genomic DNA
>  my question is
>1 Whether it can be cloned in TA cloning vector the required size.
>2. how to calculate the concentration of the PCR product after the
>reaction
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Shifali Chatrath
Graduate Student
Protein science Lab
Dept. of Biological sciences
National University of Singapore
Singapore
+65-65161210
+65-96393449