When interpreting qRT-PCR results, which results are acceptable as fold changes, which are acceptable for housekeeping genes? e.j. a values of 0.8 raw fold changes for a "housekeeping" gene is acceptable as such? and conversely which is the threshold to say that there was "fold change"?

  • $\begingroup$ What do you mean by "raw fold"? Values not normalized to the normalization gene? $\endgroup$ – Chris Mar 26 '14 at 19:12
  • $\begingroup$ yes, values not normalised to any housekeeping $\endgroup$ – Katz Mar 26 '14 at 21:15
  • $\begingroup$ Why not? This will make comparisions hard to impossible. $\endgroup$ – Chris Mar 26 '14 at 21:21
  • $\begingroup$ please clarify, what are you answering to $\endgroup$ – Katz Mar 26 '14 at 21:31
  • $\begingroup$ Why didn't you normalize your genes? $\endgroup$ – Chris Mar 26 '14 at 21:46

As I mentioned in your previous post. If your reference gene undergoes changes then replace it with another reference and if even that doesn't work out then use a spike in. You must normalize. Interpreting the raw values can be erroneous- there could be a loading bias. Don't think much- repeating RNA isolation and/or qPCR is not a very hard job. And as Chris said, use two references if you can.

BTW, if you are fine with it then can you tell what is the gene that you are studying and what is the treatment

w.r.t what is a significant fold change.

There is no hard and fast rule to determine what is an acceptable fold change. In some robust systems a minor perturbation in the gene expression will not translate to any great biological outcome whereas there can be sensitive systems in which minor changes can affect the cell state. These things depend on how the gene is integrated in the regulatory network.

|improve this answer|||||

Your Answer

By clicking “Post Your Answer”, you agree to our terms of service, privacy policy and cookie policy

Not the answer you're looking for? Browse other questions tagged or ask your own question.