minimum length for the gene to be amplified in PCR (View forum version)



roshanbernard

Posted 12 April 2009 - 04:52 PM

hi,
i have a gene which is 32 bp...can i use the pcr and produce the multiple copies it or is it too small for the PCR to do it...wat is the minimum gene length for the PCR....

HomeBrew

Posted 12 April 2009 - 05:15 PM

Why not buy two complementary primers that cover the gene in its entirety, and also leave appropriate overhangs so you can clone it? Mix the primers, melt, and cool slowly, then ligate this now-double-stranded DNA into an appropriately digested vector.

phage434

Posted 12 April 2009 - 06:56 PM

When you do this, don't forget to either order the oligos with 5' phosporylation, or kinase the annealed product to add 5' phosphates. They won't ligate with one or the other.

T C

Posted 12 April 2009 - 07:24 PM

Hey,

The lowest I have gone is 100 bp but why don't you just order a commercial olgo (in yr case gene) of 32 bp.

Best,
TC

hi,
i have a gene which is 32 bp...can i use the pcr and produce the multiple copies it or is it too small for the PCR to do it...wat is the minimum gene length for the PCR....


HomeBrew

Posted 12 April 2009 - 07:41 PM

phage434 is right -- I forgot to mention that you must have the oligos synthesized with 5' phosphorylation.

hanming86

Posted 13 April 2009 - 03:58 AM

32bp .. just do a gene synthesis. they have this service nowadays. they synthesize it and send it in a plasmid for ya.

roshanbernard

Posted 13 April 2009 - 04:44 AM

thanks a lot everybody...i think i will order for the gene with 5' phosphorylation,....