Semi-quantitative RTPCR

xudong huang xudong.huang at
Fri Apr 11 04:48:51 EST 1997

Hi, Helen

If you put together your two sets of primers and do pcr in one reaction,
then you can express the result of your target using the B2m as the
reference, because the tube-tube differences are normalized in this way.

Hope it help.

Helen Haase wrote:
> Hi!
> I would like to know more about validating my semiquantitative RTPCR
> system.  From the same RT sample I am doing two PCR reactions, one
> using primers for the message of interest and one for
> B2-microglobulin, which should act as a control.  The PCR reactions
> are done under the same conditions, at the same time.  I have
> established that the cycle number has been limited to ensure that
> amplification is still exponential.
> Is it valid then to compare the intensity of staining of the ethidium
> bromide stained bands and express results relative to the B2M control?
> Any references on principles involved would be appreciated
> Thank you
> Helen

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