ADAR expression in Pichia pastoris uses the methanol-inducible alcohol oxidase AOX1 promoter for induction. A Zeocin resistance gene on the plasmid is used to select high copy number tandem integrations of the plasmid constructs. Preparation of extracts by grinding cultures in liquid nitrogen and purification protocols using 6 � HIS and FLAG epitope tags are described. Procedures for preparing radiolabeled dsRNA and for assaying the non-specific RNA editing activity of ADARs are described.

ADARs produced in Escherichia coli are not enzymatically active. We describe expression of the ADAR dsRNA binding domains in E. coli using current versions of the T7 promoter based Studier vectors as well as the purification of the domains.