Cloning GC Rich Linker - PCR (May/10/2006 )

Hi,

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?

Cheers

Lucy

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You could buy the two oligo strands of your DNA and then anneal them.

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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

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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)

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