In vitro enzymatic synthesis of RNA is performed by in vitro transcription or by ligation. A typical in vitro transcription technique makes use of recombinant T7 RNA polymerase. Double stranded DNA templates such as PCR amplicons or linearized plasmid DNA contain the T7 promotor sequence upstream of the desired RNA sequence.
More sophisticated RNA molecules, containing e.g. fluorophores or naturally occurring modifications, can be composed by joining different RNA fragments. The latter are obtained by chemical synthesis, transcription, or from natural RNA. Via hybridization, a complementary DNA strand organizes the correct order of the fragments, which are then joined by RNA ligase.
