Jump to content

  • Log in with Facebook Log in with Twitter Log in with Windows Live Log In with Google      Sign In   
  • Create Account

Submit your paper to J Biol Methods today!
Photo
- - - - -

polymerase for BSP


  • Please log in to reply
2 replies to this topic

#1 SimonT

SimonT

    member

  • Active Members
  • Pip
  • 4 posts
0
Neutral

Posted 23 April 2009 - 05:20 AM

Hi!

I red on this forum that polymerase for bisulfite PCR shuld be non proofreading. I want to ask is it true and if it is, why?

Which polymerase do you recommend?

Regards!

Simon

#2 methylnick

methylnick

    Veteran

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 245 posts
4
Neutral

Posted 23 April 2009 - 02:31 PM

Bisulphite conversion changes all unmethylated C's to U's. And therefore generate non-complementary strands

I believe the proof reading polymerase would either stall or try to correct the mismatches arising from this base change in the first few cycles. This could potentially overestimate or underestimate the methylation calls as you are trying to detect a C or T for DNA methylation.

Nick

#3 phage434

phage434

    Veteran

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 2,484 posts
251
Excellent

Posted 08 May 2009 - 05:56 PM

The PCR enzyme is only copying a single strand -- the strands are denatured to single stranded. The problem with proof reading enzymes is the uracil, which poisons the proof reading enzyme copying reaction by stalling the enyzme.




Home - About - Terms of Service - Privacy - Contact Us

©1999-2013 Protocol Online, All rights reserved.