Identification and characteristics of microRNAs from Bombyx mori.

BACKGROUND: MicroRNAs (miRNAs) are small RNA molecules that regulate gene expression by targeting messenger RNAs (mRNAs) and causing mRNA cleavage or translation blockage. Of the 355 Arthropod miRNAs that have been identified, only 21 are B. mori miRNAs that were predicted computationally; of these, only let-7 has been confirmed by Northern blotting. RESULTS: Combining a computational method based on sequence homology searches with experimental identification based on microarray assays and Northern blotting, we identified 46 miRNAs, an additional 21 plausible miRNAs, and a novel small RNA in B. mori. The latter, bmo-miR-100-like, was identified using the known miRNA aga-miR-100 as a probe; bmo-miR-100-like was detected by microarray assay and Northern blotting, but its precursor sequences did not fold into a hairpin structure. Among these identified miRNAs, we found 12 pairs of miRNAs and miRNA*s. Northern blotting revealed that some B. mori miRNA genes were expressed only during specific stages, indicating that B. mori miRNA genes (e.g., bmo-miR-277) have developmentally regulated patterns of expression. We identified two miRNA gene clusters in the B. mori genome. bmo-miR-2b, which is found in the gene cluster bmo-miR-2a-1/bmo-miR-2a-1*/bmo-miR-2a-2/bmo-miR-2b/bmo-miR-13a*/bmo-miR-13b, encodes a newly identified member of the mir-2 family. Moreover, we found that methylation can increase the sensitivity of a DNA probe used to detect a miRNA by Northern blotting. Functional analysis revealed that 11 miRNAs may regulate 13 B. mori orthologs of the 25 known Drosophila miRNA-targeted genes according to the functional conservation. We predicted the binding sites on the 1671 3'UTR of B. mori genes; 547 targeted genes, including 986 target sites, were predicted. Of these target sites, 338 had perfect base pairing to the seed region of 43 miRNAs. From the predicted genes, 61 genes, each of them with multiple predicted target sites, should be considered excellent candidates for future functional studies. Biological classification of predicted miRNA targets showed that "binding", "catalytic activity" and "physiological process" were over-represented for the predicted genes. CONCLUSION: Combining computational predictions with microarray assays, we identified 46 B. mori miRNAs, 13 of which were miRNA*s. We identified a novel small RNA and 21 plausible B. mori miRNAs that could not be located in the available B. mori genome, but which could be detected by microarray. Thirteen and 547 target genes were predicted according to the functional conservation and binding sites, respectively. Identification of miRNAs in B. mori, particularly those that are developmentally regulated, provides a foundation for subsequent functional studies.

Analysis of similarity/dissimilarity of DNA sequences based on a class of 2D graphical representation.

On the basis of a class of 2D graphical representations of DNA sequences, sensitivity analysis has been performed, showing the high-capability of the proposed representations to take into account small modifications of the DNA sequences. And sensitivity analysis also indicates that the absolute differences of the leading eigenvalues of the L/L matrices associated with DNA increase with the increase of the number of the base mutations. Besides, we conclude that the similarity analysis method based on the correlation angles can better eliminate the effects of the lengths of DNA sequences if compared with the method using the Euclidean distances. As application, the examination of similarities/dissimilarities among the coding sequences of the first exon of beta-globin gene of different species has been performed by our method, and the reasonable results verify the validity of our method.