Abstract
BackgroundMicroRNA (miRNA) and other small regulatory RNAs contribute to the modulation of a large number of cellular processes. We sequenced three small RNA libraries prepared from the whole body, and the anterior-middle and posterior silk glands of Bombyx mori, with a view to expanding the repertoire of silkworm miRNAs and exploring transcriptional differences in miRNAs between segments of the silk gland.ResultsWith the aid of large-scale Solexa sequencing technology, we validated 257 unique miRNA genes, including 202 novel and 55 previously reported genes, corresponding to 324 loci in the silkworm genome. Over 30 known silkworm miRNAs were further corrected in their sequence constitutes and length. A number of reads originated from the loop regions of the precursors of two previously reported miRNAs (bmo-miR-1920 and miR-1921). Interestingly, the majority of the newly identified miRNAs were silkworm-specific, 23 unique miRNAs were widely conserved from invertebrates to vertebrates, 13 unique miRNAs were limited to invertebrates, and 32 were confined to insects. We identified 24 closely positioned clusters and 45 paralogs of miRNAs in the silkworm genome. However, sequence tags showed that paralogs or clusters were not prerequisites for coordinated transcription and accumulation. The majority of silkworm-specific miRNAs were located in transposable elements, and displayed significant differences in abundance between the anterior-middle and posterior silk gland.ConclusionsConservative analysis revealed that miRNAs can serve as phylogenetic markers and function in evolutionary signaling. The newly identified miRNAs greatly enrich the repertoire of insect miRNAs, and provide insights into miRNA evolution, biogenesis, and expression in insects. The differential expression of miRNAs in the anterior-middle and posterior silk glands supports their involvement as new levels in the regulation of the silkworm silk gland.
Highlights
MicroRNA and other small regulatory RNAs contribute to the modulation of a large number of cellular processes
To extend the known repertoire of small regulatory RNAs expressed in the silkworm, we constructed and sequenced three small RNA libraries prepared from the whole body (WB) as well as the anterior-middle and posterior silk glands (AMSG and PSG) of day-3 fifth instar larvae
Overall complexity of small RNA pools between the libraries We obtained raw data by sequencing three small RNA pools of the whole silkworm body from 5th-instar day-3 larvae, and anterior-middle and posterior silkworm silk glands, using the latest sequencing Solexa technology [8,20], filtered the low quality reads according to base quality value, trimmed the adaptor sequence at the 3’ primer terminus, cleaned up 5’ adaptor contaminants formed by ligation, and collected the small RNAs and analyzed size distribution
Summary
MicroRNA (miRNA) and other small regulatory RNAs contribute to the modulation of a large number of cellular processes. We sequenced three small RNA libraries prepared from the whole body, and the anteriormiddle and posterior silk glands of Bombyx mori, with a view to expanding the repertoire of silkworm miRNAs and exploring transcriptional differences in miRNAs between segments of the silk gland. Following their initial discovery in worms, an increasing number of 18-30 nt-sized small RNAs have been identified as crucial regulatory molecules in multicellular organisms, animal viruses, and unicellular organisms [1,2,3,4,5,6,7]. The newly identified miRNAs significantly enhance our knowledge of insect miRNA species and provide insights into miRNA evolution, biogenesis, and expression in insects
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