Home-made PCR gems

Basavareju Shankarappa bsh at MED.PITT.EDU
Mon Jul 20 09:16:08 EST 1992


> 
> In article <01GMHDD8X6MS000A02 at irri.cgnet.com> IYAP at IRRI.CGNET.COM writes: > >Our lab happened to have several bags of Paraplast (tissue embedding > >medium) lying around, so someone decided to test this.  He got
> >great results.  Ordinary paraffin works good too, but the Paraplast
> >is already in the form of pellets ready for use.  It also has
> >DMSO, which probably helps.
> >
> >Noel Yap
> >(Usual disclaimers apply)
> 
> I have found that the addition of DMSO generally increases the yeild 
> of a PCR reaction, as well as the specificity. I do it in 10% DMSO final
> concentration (high quality spectragrade, kept in aliquots at -80).
> Previously, it would take 40 cycles to get my product. With DMSO,
> a good band appears in only 20.
> 
> Well, PCR Gems help too :)

One word of caution to all those who would like to use DMSO and GLYCEROL.  
DMSO and nominal concentrations of GLYCEROL are known to reduce the     
effect of mismatches on primer binding.  So if specificity in amplification
is important, I believe you should avoid using these agents at least in the
initial rounds of amplification. 
So in effect although you are getting the benefits of hot-start PCR, your
specificity might be reduced.

Raj Shankarappa.
 



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