New England Biolabs, E2040S, HiScribe® T7 High Yield RNA Synthesis Kit

Now includes separate tube of DTT Related Categories RNA Capping,, sgRNA Synthesis,, RNA Synthesis In vitro Transcription (IVT), Specification Materials Required but not Supplied Nuclease-free Water (NEB #B1500) FAQ Q: Can I use the Monarch Spin RNA Cleanup Kits to cleanup my in vitro transcription (IVT) reaction? A: Yes, the Monarch Spin RNA Cleanup Kit (50 µg) (NEB #T2040) is suitable for cleaning up in vitro transcription reactions where the yield is not expected to exceed 50 µg. If the RNA yield of an in vitro transcription is expected to exceed 50 µg, we recommend the reaction be split among multiple Monarch Spin RNA Cleanup Columns (50 µg) or cleaned up using the Monarch Spin RNA Cleanup Kit (500 µg) (NEB #T2050), due to the larger RNA binding capacity. Q: How can I improve on a low yield of RNA from the transcription reaction? A: If the transcription reaction with your template generates full-length RNA but the yield is significantly lower than expected, it is possible that contaminants in the DNA template are inhibiting the RNA polymerase, or the DNA concentration may be incorrect. Alternatively, additional purification of DNA template may be required. Adding 5mM DTT (final) to the transcription reaction may also help improve RNA yield. Q: Are modified nucleotides included in the kit? A: Modified nucleotides are not included in the kit but can be purchased separately. NEB can provide N1-Methyl-Pseudouridine-5’-Triphosphate (NEB #N0431), 5-Methyl-Cytidine-5’-Triphosphate (NEB #N0432), Pseudouridine-5’-Triphosphate (NEB #N0433) and 5-Methoxy-Uridine-5’-Triphosphate (NEB #N0434). The kit manual includes a detailed protocol for using modified nucleotides in the transcription reaction. Q: Do I need to add DTT to the reaction? A: Addition of DTT (5mM final) to the reaction is optional but recommended.The RNA polymerase in the kit is sensitive to oxidation and could result in lower RNA yield over time due to repeated handling etc. Adding DTT to the reaction may help restore the kit performance in such cases. Adding DTT will not compromise the reaction in any situation.