Cotranscriptional 3′-End Processing of T7 RNA Polymerase Transcripts by a Smaller HDV Ribozyme
In vitro run-off transcription by T7 RNA polymerase generates heterogeneous 3′-ends because the enzyme tends to add untemplated adenylates. To generate homogeneous 3′-termini, HDV ribozymes have been used widely. Their sequences are added to the 3′-terminus such that co-transcriptional self-cleavage generates homogeneous 3′-ends. A shorter HDV sequence that cleaves itself efficiently would be advantageous. Here we show that a recently discovered, small HDV ribozyme is a good alternative to the previously used HDV ribozyme. The new HDV ribozyme is more efficient in some sequence contexts, and less efficient in other sequence contexts than the previously used HDV ribozyme. The smaller size makes the new HDV ribozyme a good alternative for transcript 3′-end processing.
KeywordsIn vitro transcription HDV ribozyme Co-transcriptional processing 3′-Terminus
We thank Andrej Luptak for helpful discussions. Partial support to U.F.M came from grant NASA NNX13AJ09G and NASA grant NNX16AJ27G. Support for A.A. was provided by a 2015/2016 Department of Education Graduate Assistance in Areas of National Need (GAANN) Training Grant # P200A150251, and by a 2016 Distinguished Graduate Student Fellowship by the Department of Chemistry & Biochemistry at UC San Diego.
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