In Vitro Transcription on Zageno
The in vitro transcription process
The starting template of in vitro transcription is a linear DNA template that includes a promoter region together with the usual ingredients for a successful transcription process: RNA polymerase, nucleotides, and an appropriate buffer system. Since every reaction is different, the conditions may vary depending on your DNA template, expected RNA yield, length, and purity.
Make use of our In Vitro Transcription Troubleshoot for any experimental issues.
You need a double stranded DNA (dsDNA) template, but its origin is irrelevant. The template can be anything from normal PCR reactions, plasmids, or even cDNA templates that themselves were generated from RNA molecules. However, different kits may be suitable for different templates, so take care when choosing your kit.
- In order to use a PCR product in an in vitro transcription reaction, you have to add a promoter to your fragment so that it is recognized by RNA polymerase. The promoter is added at the 5’ end of the fragment.
- Plasmid vectors must first be linearized before the transcription reaction can ensue. This is (fairly) easily done by using restriction enzymes. Purification should be carried out after such a digest as the restriction enzymes may inhibit the downstream in vitro transcription process.
- One of the latest developments in this field is using in vitro transcription in RNA amplification reactions. Here, the reaction template is created from RNA during reverse transcription, generating cDNA. A second strand synthesis reaction is then performed to convert the cDNA to dsDNA for the in vitro transcription to be carried out.
Other applications also exists such as the use of oligonucleotides as templates.
Compare In Vitro Transcription Kits
With our compare function, you can avoid all the time and energy wasted sifting through multiple web pages from different suppliers. At ZAGENO you can clearly see kits side-by-side, with the relevant attributes for each kit neatly in line for easy selection of the best product for you.
Click on the comparison below for a clearer view!
For example, this comparison shows:
- that the SP6/T7 TRANSCRIPTION Kit from Roche Life Science is suitable for downstream in vitro and in vivo translation.
- that the MEGASCRIPT® SP6 TRANSCRIPTION Kit from Invitrogen is the cheapest per reaction.
- that the RIBOPROBE® COMBINATION SYSTEM-SP6/T7 RNA POLYMERASE Kit only requires an hour to produce the final sample.
This comparison clearly exemplifies how every kit has strengths and weaknesses. Depending on what features you require for your experiment, these details should enable you to make an informed decision on the right kit for you.
The ZAGENO comparison does not highlight one kit to be better than the other, as the kit of choice may vary between researchers - depending on each individual's preferred attributes. The best kit is the one that meets your needs - ZAGENO allows you to make an informed decision with minimum effort.
Check out our How It Works page for a guide to using the comparison function.