Functional messenger RNAs are produced by SP6<i>in vitro</i>transcription of cloned cDNAs

TLDR

The study presents a method to synthesize microgram quantities of eukaryotic mRNAs. The approach uses in vitro transcription of cDNA clones inserted into SP6 vectors to produce mRNA. Synthetic mRNAs produced by this method function efficiently in frog oocyte cytoplasm and wheat germ extracts, require a 5′ cap for translation, tolerate deletion of most 3′ flanking sequences, and enable large‑scale production of protein from any cDNA clone.

Abstract

We describe a method for the synthesis of microgram quantities of eucaryotic messenger RNAs. Injection into the cytoplasm of frog oocytes and addition to wheat germ extracts show that these synthetic RNAs function efficiently as messenger RNAs. We confirm that a 5′ cap on the mRNA is essential for translation in injected oocytes and show that most of the 3′ flanking region, including the poly A tail, can be deleted without the abolition of protein synthesis. The method of mRNA synthesis involves invitro transcription of cDNAs which have been cloned into SP6 vectors (described in the accompanying paper). This method enables one to produce large amounts of mRNA and consequently protein from any cDNA clone.