AmpliScribe™ T7, T3 and SP6 High Yield Transcription Kits from Epicentre Biotechnologies

In vitro transcription technology is high performance technique which enables researchers to effectively study the RNA world. Hence a tool that generates high yields of RNA in vitro is welcome. In a standard reaction, the AmpliScribe™ T7, T3 and SP6 High Yield Transcription Kits generate tens of micrograms of the desired RNA within two hours, starting from just 1 microgram of DNA template. The kits contain everything needed for the transcription reaction: separate solutions of appropriate nucleotides, the specific enzyme T7, T3 or SP6, dithiothreitol (DTT), reaction buffer, DNase I, RNase-free water and, in addition, a linearized plasmid is included for use as a control template. The standard protocol is very easy and the reaction can be set up at room temperature, avoiding the cold-related distress of the researcher. Once the tube is prepared by adding the reaction components, you have just to put it to 37°C for 2 hours. Then, DNA digestion and RNA purification steps can be carried out, if desired.

Although these kits are not dedicated to the production of labeled RNA, through the modification of the standard protocol, the kit can synthesize fluorescent or non-isotopic digoxigenin (DIG) (available separately from Roche) labelled RNA. (Note: The high yield kits are affordably priced in comparison with standard non-isotopic labelling kits, e.g. DIG-labelling RNA kit from Roche.) The protocol can be also modified to produce 5'-capped RNA (cap analogs are available separately).

I used the AmpliScribe T7, T3 and SP6 High Yield Transcription Kits to produce large amounts of DIG-labeled RNA probes using digoxigenin 11-UTP (Roche) following the modified protocol of Kaplan and colleagues (available from Epicentre Biotechnologies (1)). I produced three sense and three antisense DIG RNA probes and I found that the DIG labeling reaction yields generally less than the standard protocol; however, I generated enough to perform in situ hybridization on almost 100 sections for each probe.

Overall, the AmpliScribe T7, T3 and SP6 High Yield Transcription Kits produce high yields of RNA and are the tools of choice to carry out in vitro RNA studies.

Reference
(1) Kaplan, E.D. et al., (1996) EPICENTRE Forum, 3 (2), 1.