Identification and characterization of new miRNAs cloned from normal mouse mammary gland

To extend the repertoire of mouse mammary gland expressed miRNAs, we have constructed several libraries of small miRNAs allowing the cloning of 455 sequences. After bioinformatics' analysis, 3 known miRNA (present in miRbase) and 33 new miRNAs were identified. Expression of 24 out of the 33 has been confirmed by RT-PCR. Expression of none of them was found to be mammary specific, despite a tissue-restricted distribution of some of them. No correlation could be established between their expression pattern and evolutionary conservation. Six of them appear to be mouse specific. In several cases, multiple potential precursors of miRNA were present in the genome and we have developed a strategy to determine which of them was able to mature the miRNA.The cloning approach has allowed improving the repertoire of miRNAs in the mammary gland, an evolutionary recent organ. This tissue is a good candidate to find tissue-specific miRNAs and to detect miRNA specific to mammals. We provide evidence for 24 new miRNA. If none of them is mammary gland specific, a few of them are not ubiquitously expressed. For the first time 6 mouse specific miRNA have been identified.Numerous small non-coding RNAs of 18–25 bases in length, called microRNAs (miRNAs), have been found to play important roles in silencing specific target genes. Recently Vasudevan et al. [1] have shown that miRNAs can also activate gene expression, inducing translation up-regulation of target messager RNAs (mRNAs) on cell cycle arrest. The total estimated number of reasonably conserved miRNAs in vertebrates varies from 250 [2] to 600 [3]. In human, Bentwich et al. [4] suggested that the total number of miRNAs is above 800. The sequences of many miRNAs are conserved among distantly related organisms [5], but recent evidences demonstrated the presence of primate-specific miRNAs [6,7]. miRNAs are transcripts which are cleaved from a ~70 nucleotides hairpin precursor by Dicer [8,9]. They regulate gene expression at the postt