Design and in vitro transcription of shRNA with theNEB HiScribe™ T7 Quick High Yield RNA Synthesis Kit v1

This a protocol for design and in vitro transcription (IVT) of shRNA with T7 RNA polymerase from a dsDNA template created by annealing two complementary ssDNA oligos. A single reaction should produce between 30 and 150 µg shRNA depending on the sequence directly downstream of the T7 promoter (and to some part probably the overall sequence). The protocol was used to produce shRNA to target different mRNAs for RNAi-based knockdown after electroporation into Aiptasia zygotes.