Analysis of miR-30 expression level. (A) A schematic of pCMV-miR-30. The predicted miR-30 precursor (Table 1) is flanked by XhoI sites, and the basal stem is, therefore, predicted to be extended by 6 bp. PA site, polyadenylation site. (B) Primer extension assay to detect miR-30 in transfected 293T cells. Lane 1, RNA from cells transfected with pBC12/CMV; lane 2, pCMV-miR-30; lanes 3–7, various miR-30 mutants. (C) Northern analysis of miR-30 expression. Lane 1, pBC12/CMV; lane 2, pCMV-miR-30; lanes 3–5, various miR-30 mutants. The arrow indicates the predicted position of the mature miR-30 and the arrowhead indicates the ∼65-nt mir-30 pre-miRNA on the Northern blot. Positions of known DNA markers are shown at left.
