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
- - - - -

Full length cDNA


  • Please log in to reply
4 replies to this topic

#1 noyara

noyara

    Enthusiast

  • Active Members
  • PipPipPipPipPip
  • 59 posts
1
Neutral

Posted 14 September 2011 - 05:11 AM

Hi
How to get full length cDNA by using PCR? in the normal way?
I try to design primers in the end of desire gene but still there are some base pares out of primer sequence,
and if i try to make the primer start from the first base but the quality of it will not be acceptable

Do you have any suggestion..

Thank you in advance
Noor

#2 MurphysunHKU

MurphysunHKU

    member

  • Active Members
  • Pip
  • 24 posts
0
Neutral

Posted 14 September 2011 - 04:31 PM

U can design a pair of primers that bind to the vector specific sequence that flank your insert site. And after PCR , u can use this as a probe to rescreen your cDNA library to increase the chance of having a full length cDNA clone

#3 bob1

bob1

    Thelymitra pulchella

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 5,674 posts
396
Excellent

Posted 14 September 2011 - 09:49 PM

Check out a protocol known as RACE - Rapid Amplification of cDNA Ends.

#4 noyara

noyara

    Enthusiast

  • Active Members
  • PipPipPipPipPip
  • 59 posts
1
Neutral

Posted 14 September 2011 - 10:15 PM

thank you MurphysunHKU for your reply,


Hi bob1,
I think this kit is not cheap, also can run 10 reactions only.. Is it?

#5 bob1

bob1

    Thelymitra pulchella

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 5,674 posts
396
Excellent

Posted 15 September 2011 - 09:03 PM

Kits aren't cheap, but buying the components and looking up the protocols is relatively cheap. Basically the kits are just a PCR kit with primers and maybe a terminal transferase. There are two sorts of RACE, 5' and 3'. 3' is much easier than the 5', and wikipedia describes the basic process and differences reasonably well.




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

©1999-2013 Protocol Online, All rights reserved.