Validation of predicted miRNAs

The processing of primary transcripts ectopically expressed in S2 cells under the control of the actin 5C promoter was analyzed by Northern blot using probes designed against both arms of the predicted hairpins. In 16 cases, mature sequences could be detected (column 6, Y), five of which were present in untransfected S2 cells. For the remaining predictions, no processing was observed (column 6, N). In most cases, small RNAs corresponding to the probed sequences could also be detected in published small RNA libraries (13/16). Six miRNAs either not assayed (colum 5, n.a.) or not detected by Northern blot could be validated, too. No matching sequences were found in the libraries for locus7, locus25 and manual3, which can be detected as endogenous transcripts (column 6, endogenous) by Northern blot.