quantification of RNA

nienke vriezen n.vriezen at stm.tudelft.nl
Tue Jun 18 08:43:50 EST 1996


The orcinol method has worked for me in determining the RNA content of 
mammalian cells. A 1 ml sample ( in my case a HCLO4 RNA precipitate )
was pipetted into a glas tube large enough and strong enough to stand the 
rest of the protocol. To this 3 ml of the orcinol mixture was added ( 4 
parts 90 mg FeCl3.6H2O in 100 ml HCL and 1 part 1gr orcinol in 100 ml H2O 
). This mixture is heated to 100 C for 20 minutes, then cooled with tap 
water. 8 to 10 ml of butanol is added and the A672 nm is read.
You can use ribose as a standard, then you have to time the 20 minutes at 
100 C exactly, if you use RNA as a standard this isn't as critical.
As the method is based on a reaction with the sugar other sugars may 
interfere.
A good reference is Methods in microbiology part 5B, Eds Norris,J.R.; 
Ribbons, D.W. Acad.Press. '71 London, New York.

good luck 
Nienke




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