CyPhyRNA-seq: a genome-scale RNA-seq method to detect active self-cleaving ribozymes by capturing RNAs with 2',3' cyclic phosphates and 5' hydroxyl ends.

ABSTRACT: Self-cleaving ribozymes are catalytically active RNAs that cleave themselves into a 5'-fragment with a 2',3'-cyclic phosphate and a 3'-fragment with a 5'-hydroxyl. They are widely applied for the construction of synthetic RNA devices and RNA-based therapeutics. However, the targeted discovery of self-cleaving ribozymes remains a major challenge. We developed a transcriptome-wide method, called cyPhyRNA-seq, to screen for ribozyme cleavage fragments in total RNA extract. This approach employs the specific ligation-based capture of ribozyme 5'-fragments using a variant of the Arabidopsis thaliana tRNA ligase we engineered. To capture ribozyme 3'-fragments, they are enriched from total RNA by enzymatic treatments. We optimized and enhanced the individual steps of cyPhyRNA-seq in vit

SUBMITTER: Olzog VJ 

PROVIDER: S-EPMC8782182 | biostudies-literature | 2021 Nov

REPOSITORIES: biostudies-literature