Novel RNA molecular bioengineering technology efficiently produces functional miRNA agents

Heterologous overexpression of a total of 48 new BioRNAs with human glycyl or leucyl tRNA fused hsa-pre-miR-34a carrier (BioRNA^Gly or BioRNA^Leu). (A) The 2D graphic of BioRNA^Gly or BioRNA^Leu with a payload miRNA whose guide and passenger sequences are highlighted. (B) Sequence design of target BioRNA inserts for cloning into a pBSKrna vector linearized by EcoRI and PstI restriction enzymes to offer the BioRNA expression plasmid (pBSK/BioRNA), which is comprised of a lipoprotein (lpp) promoter, terminator from the Escherichia coli ribosomal RNA rrnC operon (rrnC), and the gene encoding β-lactamase (bla) for antibiotic resistance and selection. (C, D) Urea–PAGE analyses depicting the successful overexpression of 26 BioRNA^Gly (C) and 22 BioRNA^Leu (D) versions of recombinant miRNAs, as manifested by the appearance of a strong new band at the expected size when compared to total RNA from the wild-type HST08 E. coli; 300 ng RNA per lane. Urea–PAGE analyses depicting the successful overexpression of BioRNA^Leu/miR-126b-3p, -133a-3p, -200b-3p, and -7-5p have been reported recently (Li et al. 2021).