Help RT-PCR!

George W Chacko gchacko at magnus.acs.ohio-state.edu
Tue Nov 5 15:09:18 EST 1991


I work with human platelet RNA and as one of the negative controls for RT-PCR
in this system I need a no RT enzyme tube to ensure that I don't have 
contaminating DNA in my sample. 

I've been working with the Pharmacia cDNA synthesis kit which has provided
good results so far except that they supply what they call a Bulk First
Strand Reaction Mixture reagent which is a cocktail of dNTP's, buffer,
salt and MMLV-RT.

The alternative is to inactivate the RT completely which leads me in long
winded rambling way to my question. What's the best way to inactivate
MMLV-RT? I thought of boiling for 10 minutes and Proteinase K treatment
followed by heat inactivation.

All suggestion and comments are greatly appreciated.

Thanks

George



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