Rapid preparation of M1RNA subunit of ribozyme P in vitro

Objective To prepare the core catalytic subunit M1RNA of prokaryotic source ribozyme P rapidly. Methods Using Escherichia coli DH5 as the template, the M1RNA gene was amplified by PCR and placed in the downstream of T7 promoter. The PCR products were identified by electrophoresis and sequencing. Using the M1RNA gene as the template and 32P labelling, M1RNA was obtained by the action of T7 RNA polymerase. The products were separated by urea denaturing polyacrylamide gel and the results were observed. Results PCR method was used to amplify the M1RNA gene from the Escherichia coli genome, and the M1RNA gene was successfully amplified in vitro and identified by gel electrophoresis and sequencing. The M1RNA was placed in the downstream of the T7 promoter. Under the action of T7RNA polymerase, the products were isolated by urea denaturing polyacrylamide gel, and the results showed that the band size was 377 nt, which is consistent with the expected size. These confirmed that M1RNA was successfully obtained in vitro. Conclusion The independent catalytic subunit of M1RNA has been successfully prepared rapidly in vitro, and the gene silencing technology based on this has good prospects for developing new antisense nucleic acid drugs. Key words: Ribozyme P; M1RNA; Gene silencing; Transcription