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
- - - - -

TDNA Genotyping PCR program


  • Please log in to reply
1 reply to this topic

#1 praveen921

praveen921

    member

  • Active Members
  • Pip
  • 7 posts
0
Neutral

Posted 05 October 2012 - 02:47 AM

Hello all,

I am in search for a PCR program for screening TDNA inserts in plants.The annealing temp of my Primer set is 61C.Could someone help me with the PCR steps i have to use for TDNA genotyping in plants?

Thanks in advance.

#2 bob1

bob1

    Thelymitra pulchella

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 5,738 posts
400
Excellent

Posted 05 October 2012 - 01:45 PM

Your best bet will be to design the PCR program for yourself, as the exact conditions depend on the primers you use, the PCR machine (ramp times, stability), and the polymerase used.

A basic PCR program is this:
1)95 for 5 min,
2)95 for 1 min
3)Primer annealing temp for 1 min (optimise this for your primers. Note: Tm (melting temp) is not the same as Ta (annealing temp)
4)72 for 1 min)
5)repeat steps 2, 3, 4, 30x
6)72 for 5 min.

The exact extension time to use is dependent on the processivity of the polymerase, Taq will do about 1000 bp/10s.




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

©1999-2013 Protocol Online, All rights reserved.