A general method for rapid and cost-efficient large-scale production of 5' capped RNA.

ABSTRACT: The eukaryotic mRNA 5' cap structure is indispensible for pre-mRNA processing, mRNA export, translation initiation, and mRNA stability. Despite this importance, structural and biophysical studies that involve capped RNA are challenging and rare due to the lack of a general method to prepare mRNA in sufficient quantities. Here, we show that the vaccinia capping enzyme can be used to produce capped RNA in the amounts that are required for large-scale structural studies. We have therefore designed an efficient expression and purification protocol for the vaccinia capping enzyme. Using this approach, the reaction scale can be increased in a cost-efficient manner, where the yields of the capped RNA solely depend on the amount of available uncapped RNA target. Using a large number of RNA substra

SUBMITTER: Fuchs AL 

PROVIDER: S-EPMC4986899 | biostudies-literature | 2016 Sep

REPOSITORIES: biostudies-literature