I have noticed whilst using Beta 2 Microglobulin as a reference gene, the expression reduced significantly whilst doing PCR, can any one explain this? Cells were stimulated with LPS.
try normalization with another reference gene, as it could be instable...see for example: http://rd.springer.com/article/10.1186%2F1471-2172-11-21
and it is Real-time PCR or end-point PCR
It's RT-PCR, I am working with macrophage cell line (RAW 264.7) and lymphocytes, the paper recommends beta 2 microglobulin, with monocytes, ( I found a similar paper, hence why I selected it as a reference) I have tested this now with B2M and found B2M to be highly unstable.
guessed the problem could be in the reference gene..unfortunately this could happen..remember to do check always for stability of the reference gene...and go for two different reference :) good luck!
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