Once your gene of interest is seamlessly cloned into the template vector, linearize the plasmid with a restriction enzyme such as HindIII (not included) to create the double-stranded DNA containing the T7 promoter sequence. The template linearization ensures IVT transcripts are produced at a uniform length.
Takara IVTpro T7 mRNA Synthesis Kit is optimized for use with cap analogs such as CleanCap Reagent AG (TriLink Biotechnologies, Cat. # N-7113, not included). Please see the Co-transcriptional capping section for details. If using any IVT template vector other than the Linearized Template Vector in the Cloning Kit for mRNA Template (Cat. # 6143), verify that it has a transcription start site compatible with the cap analogs used in the IVT reaction.
The IVT reaction run with the Takara IVTpro T7 mRNA Synthesis Kit can be scaled up 10-fold. A consistent mRNA yield across this range has been demonstrated using CleanCap Reagent (Figure 1).
Takara IVTpro T7 mRNA Synthesis Kit also contains DNase I to digest the template DNA after the IVT reaction and a lithium chloride (LiCl) solution to purify the synthesized mRNA.