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
- - - - -

Untreated samples negative, how to analyze fold change?

fold change untreated analysis

  • Please log in to reply
1 reply to this topic

#1 AnneUW

AnneUW

    member

  • Members
  • Pip
  • 4 posts
0
Neutral

Posted 29 January 2014 - 02:31 PM

I'm trying to analyze a set of data.  For one gene, the untreated samples are basically negative and are very close to the NTCs' Ct values.  The treated samples have a very good increase in expression.  Following my usual policy of not using data within 5 Ct of the negative control, I used a new primer set on the same cDNA but got a similar result.  Is there a way to use this data and phrase the analysis in a way to make it clear that the fold change is not exact since the untreated samples have such low expression?  I'm sure this happens, but I am having trouble phrasing a search to bring up published papers that have had this issue.



#2 Trof

Trof

    Brain on a stick

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 1,149 posts
101
Excellent

Posted 03 February 2014 - 03:11 AM

I've seen the way of analyzing negative data as using the "number of cycles in the program"+1 as a 'Ct' for the negative calibrator. However I don't have any reference for that (I heard it on a qPCR conference and didn't make notes) and I even think I was looking for it on several occasions.

You could mention it under the results table, how you acquired calibrator Ct for this analysis.

 

But generaly is better to use a positive sample as a calibrator, but some genes can have virtually zero expression in normal or control/untreated samples, eventhough other genes are normalized well. The aforementiond method is used exactly in this case, when you can't get a better calibrator or/and it's a part of a larger analysis set.


Our country has a serious deficiency in lighthouses. I assume the main reason is that we have no sea.

I never trust anything that can't be doubted.

'Normal' is a dryer setting. - Elizabeth Moon






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

©1999-2013 Protocol Online, All rights reserved.