In vitro transcription kits & other RNA reagents

[Kenneth Kao]

Donald Chun Kit Wong (dcwong at unixg.ubc.ca) wrote:
: We are going to begin to do some in situ hybridization in our lab and 
: need to ask for some advice.

: We are going to do in vitro transcription to make RNA probes from cDNA 
: inserted in plasmid. We have seen several kits from companies, eg. 
: Stratagene, Ambion, Boehringer Mannheim, etc. Has anyone found whether 
: one is better than the others?


We routinely make synthetic RNA for injection into cells, RNAase 
protection assays and whole mount in situ digoxigenin probes. None of the 
kits work as well as when we assemble the reactions ourselves. In fact, 
we had to abandon using Promega's Ribomax kit for RNA injection into 
embryos because we were getting non-specific effects due to insufficient
capping of messages. I can e-mail or FAX you our protocols if you are 
interested.


: We have also seen other RNA products, eg. RNaseZap, dip sticks to detect 
: RNase in solutions, etc. and in situ hybridization kits. Is there any of 
: these that are useful or recommended to be used?

: Any other suggestions / tips will be appreciated.

: Donald Wong
: Pathology, Univ. of B.C.
: Vancouver, Canada