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
- - - - -

Estimation of DNA Quantity on an Agarose Gel


  • Please log in to reply
5 replies to this topic

#1 yoshi

yoshi

    member

  • Active Members
  • Pip
  • 21 posts
0
Neutral

Posted 27 December 2010 - 10:34 PM

To estimate the DNA quantity on an agarose gel stained with EtBr, should I compare the intensity of my band of interest with the standard band of a similar size, or with the standard band of a similar intensity (assuming there is no standard band that is of similar size and intensity).

#2 pDNA

pDNA

    Veteran

  • Active Members
  • PipPipPipPipPipPipPipPipPipPip
  • 493 posts
14
Good

Posted 29 December 2010 - 08:20 AM

just use an marker with bands of different size and intensities

regards,
p

#3 yoshi

yoshi

    member

  • Active Members
  • Pip
  • 21 posts
0
Neutral

Posted 29 December 2010 - 10:39 PM

just use an marker with bands of different size and intensities

regards,
p


So with which of the bands of the marker would you recommend that I compare? The one that is of similar size to my band, or the one that is of similar intensity?

#4 scolix

scolix

    a student

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 314 posts
5
Neutral

Posted 30 December 2010 - 09:58 AM

best would be to compare similar size and intensity. But this might not be the case always. One would have to load different amounts of DNA ladder (with conc. eg. 2 log DNA - NEB) and compare intensities.

good luck

#5 hobglobin

hobglobin

    Growing old is mandatory, growing up is optional...

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 5,546 posts
104
Excellent

Posted 30 December 2010 - 12:11 PM

And be sure to use a good software. Anyway this is a suboptimal approach, though cheap, as it measures only the end-point which can differ in every reaction due to several reasons (such as deactivation of Taq, shortage of nucleotide substrates and primers, etc).
State-of-the-art an much more exact is qPCR, as the here the exponential and therefore quantifiable part of the reaction is used. But it's expensive and you need a real-time machine.
You might try a competitive PCR as alternative.

One must presume that long and short arguments contribute to the same end. - Epicurus
...except casandra's that belong to the funniest, most interesting and imaginative (or over-imaginative?) ones, I suppose.

That is....if she posts at all.


#6 yoshi

yoshi

    member

  • Active Members
  • Pip
  • 21 posts
0
Neutral

Posted 01 January 2011 - 10:34 PM

Thanks peeps!




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

©1999-2013 Protocol Online, All rights reserved.