Rarity of Novel microRNAs in the Mouse Inner Ear
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The vertebrate inner ear is a complex organ that is highly regulated throughout development by molecular and cellular factors such as morphogens and transcription factors. Another class of gene regulators whose function in the inner ear is essential but not well understood is microRNAs. Previous microarray analyses of microRNAs in the mouse inner ear performed at postnatal time-points confirmed the presence of microRNAs in the ear, suggesting that microRNAs play a role in inner ear development. To date, over 440 microRNAs have been validated in mouse. However, predictions based on bioinformatic analyses of genomic sequences suggest that the number of mammalian miRNA genes might be as great as 1000. Furthermore, most methodologies for microRNA detection only access known microRNAs. In order to gain a comprehensive knowledge of the microRNAs involved in inner ear development, microRNAs were cloned and sequenced from total RNA of embryonic day 14.5 cochlea. To increase the frequency of novel microRNA discovery, the most abundant miRNAs from an initial round of cloning and sequencing were subtracted using a novel approach, and a second round of cloning and sequencing was performed. Out of 239 sequenced isolates, only 3 unconserved novel miRNAs were found. This frequency of novel miRNA discovery suggests that the number of miRNA genes predicted by bioinformatic analyses is grossly overestimated. Further analysis demonstrates that cloned microRNAs largely represent abundant and conserved microRNAs among the many microRNAs listed for the mouse genome, suggesting that the subset of known microRNAs relevant to ear biology is substantially fewer than the total.
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