How many bases overlap in recombinant PCR?

Taek H. You tyou at postbox.acs.ohio-state.edu
Fri Oct 25 00:46:15 EST 1996


In article <54o3ls$1rb at newsy.ifm.liu.se> zywzyw at lysator.liu.se (Zhengyang Wu) writes:

>I failed to do PCR-based recombination when the overlapping was less than
>12 bp. When I made new oligoes and increased the overlapping to 18 bp, it 
>worked fine. 

>No idea to save such little amount of money: if you have to optimize the PCR
>recombination with shorter oligoes, it will cost much more than just 
>make a long one from the beginning. They also save your frustration!

While it may be true, short overlap also works.
I used 10 to 12 base overlap succefully.
However, the longer the better.
If your total length of the oligo is longer than 45 bases, PAGE purification 
would help greatly.What I noticed from non-purified oligos PCR, the band looks 
diffsed, almost looking like a local smear. It's because of the incomplete 
synthesis of the oligos.




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