FAQ: AmpliScribe™ T7-Flash and DuraScribe® T7 In Vitro Transcription Kits

  1. What is the longest in vitro transcript readily made with an AmpliScribe T7-Flash Transcription Kit?
  2. I’m using less than the recommended 1 µg of DNA template in an AmpliScribe T7-Flash reaction. How can I improve my RNA yield?
  3. Why should I set up the AmpliScribe T7-Flash reactions at room temperature? Won’t that affect the RNA polymerase enzyme activity?
  4. How clean does the template DNA need to be for in vitro transcription?
  5. What is the shortest in vitro transcript readily made with an AmpliScribe T7-Flash Transcription Kit?
  6. Can I make nonradioactive, labeled RNA using an AmpliScribe T7-Flash Kit (with labeled nucleotides or by end-labeling)?
  7. What advantages does an AmpliScribe T7-Flash Transcription Kit have over the standard AmpliScribe High-Yield Transcription Kit?
  8. Can I make radiolabeled probes with any of the AmpliScribe High-Yield or T7-Flash transcription kits?
  9. I am making a template for in vitro transcription using the AmpliScribe Kit. The template structure contains a combined double-stranded T7 promoter and a single-stranded template section. Will AmpliScribe work with this template?
  10. What is the difference between RNA made with the DuraScribe T7 Transcription Kit and RNA made with other in vitro transcription kits?
  11. My AmpliScribe reaction had precipitate while setting up the reaction. Is this normal?
  12. My AmpliScribe reaction had precipitate in the reaction tube after the protocol’s recommended 30 minutes. Did my reaction fail?
  13. What is the best procedure for cleanup/DNA template removal to recover the most RNA?
  14. What is the number of units per microliter of T7 RNA Polymerase in the AmpliScribe T7-Flash Kit?
  15. My AmpliScribe reaction had excellent yields, but when I ran it on a native gel the RNA was much shorter than the original template. What could have gone wrong?
  16. Whenever I use my AmpliScribe Kit I get a really bad smear of RNA during gel electrophoresis instead of a band that is the size of my desired transcript. What is wrong? Did an RNase contaminant get into the reaction?