Cloning GC Rich Linker - PCR (May/10/2006 )
I am attempting to clone a GC rich linker of ~60bp into a gene, which is proving very difficult!
Incorporation of the linker into primers has failed;
Does anyone have any advice how I can acheive this please?
Is it possible to buy custom dsDNA with restriction sites so that I can digest the linker and insert it into the gene directly?
You could buy the two oligo strands of your DNA and then anneal them.
Sorry to ask; but how exactly would you do that; just add the two oligos at a specific temperature for 30min maybe??? thanks for the help btw
to anneal oligos:
mix at 1:1
incubate at 95C for 5'
slowly reduce heat to 25C (I just turn off the block and let them cool over a couple hours)