Ribonuclease Protection Assay Controls ?

Martin Leach leach at acs.bu.edu
Sat Dec 17 12:28:21 EST 1994


In article <3cu257$qgu at apakabar.cc.columbia.edu>,
jjm3 at sawasdee.cc.columbia.edu (Jeffrey Joseph Martino) wrote:

> OK here it goes: Doing Ribonuclease Protection Assays on total RNA
>                  from various mouse cells/tissues (Northern no signal,
>                  and initial RPAs - no signal)
> 
>   - actin controls give a good signal.  
> 
> Actin isn't too rare.  Want to put a detection limit number on that
> RPA negative result.
> 
> I thought adding known quantities of an RNA back to sample as a control
> would be OK to check detection limit, but I promised to keep an eye out
> for an endogenous alternative:
> 
>   Any ideas for a good (not tissue-specific) low level message (with
>   probes available) for a control?
> 

Adding labelled double stranded DNA is a good loading error/processing
control.

GAPDH (metabolic enzyme) is available from clonetech...however, it does
vary with drug treatments etc...in our hands...we use b-actin/GAPDH


-- 

.....          Martin Leach                Email:leach at mbcrr.harvard.edu 
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