Unknown,Transcriptomics,Genomics,Proteomics

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Small RNA-seq analysis of single porcine blastocysts revealed that maternal estradiol-17beta exposure does not affect miRNA isoform (isomiR) expression

ABSTRACT: Small RNA sequencing of 36 porcine embryo samples collected on Day 10 of pregnancy detected miRNA sequences mapping to known and predicted porcine miRNAs, and novel miRNAs highly conserved in human and cattle. A set of highly abundant miRNAs was identified as well as a large number of rarely expressed miRNAs. Due to the high number of generated sequence reads, it was possible to detect a large number of different miRNA isoforms (isomiRs). Potentially embryo-specific miRNAs were identified by comparing the data with small RNA-Seq data sets from porcine endometrium and a search in databases for miRNA tissue expression. The identification of 140 novel miRNAs in addition to 173 known porcine miRNAs (miRBase21) showed the benefit of our small RNA analysis pipeline. This benefit has a big impact for poorly annotated species such as the pig where it is possible to annotate many additional miRNAs based on the knowledge from related species. Furthermore, we integrated our pipeline into a Galaxy workflow using standard Galaxy tools which guaranties high reproducibility and flexibility for new/upcoming datasets/studies. Using ortholog species information increases the total number of annotated miRNAs while mapping to other non-coding RNAs decreased the chances of falsely annotated miRNAs. Galaxy workflows can be published, shared, downloaded, and optimized to adapt to other species or modify analysis parameters, such as tools specifically developed for a given research project/problem, so that a broad audience can use it.

INSTRUMENT(S): Illumina HiSeq 4000

ORGANISM(S): Sus scrofa

SUBMITTER: Jochen Bick

PROVIDER: E-MTAB-6201 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Sus scrofa (pig, or swine) is one of the most important economic animals and a close biological model for complex human diseases such as obesity and diabetes. It is therefore utterly important to decode the porcine microRNAome (miRNAome) as in the literature only a small portion of it is known. In this work, a comprehensive search for porcine microRNAs (miRNAs) by Illumina sequencing was performed in ten small RNA libraries prepared from mixtures of assorted tissues, which included those collected from fetuses to adult pigs. The millions of the sequencing reads were analyzed with reference to 77 known porcine miRNA precursors (pre-miRNAs) and 3,443 distinct pre-miRNAs of other mammals listed in miRBase 13.0, and the most updated porcine genome (Sscrofa9, April 2009) and available EST sequences. Additionally, miRNA candidates specific to pig are predicated by genome & EST match and hairpin folding. Our search found 72 out of 78 (~92%) known porcine miRNAs and miRNA*s, and 36 previously unannotated miRNA*s are also indentified. Furthermore, we discovered 397 novel miRNAs by mapping to the sequencing transcripts to other mammalian pre-miRNAs and 493 candidate miRNAs which do not map to other mammalian miRNAomes and could be pig-specific. We constructed sequence- and genome-position clusters for the total of 998 miRNA candidates originating from 862 pre-miRNAs, which represent 777 unique miRNA sequences. Together with the six known porcine miRNAs that not been observed in our study, we report herein the sequence families of 783 unique miRNAs and genomic distribution patterns of 622 pre-miRNAs. We preformed q-PCR experiments for selected 30 miRNAs in 47 tissue-specific samples and found agreement between the sequencing data and the q-PCR data. We envision that our report will serve as a valuable resource for future studies aimed at understanding miRNAome of pig Ten small RNA libraries from samples of porcine fetuses (30 days after insemination) to mixed tissues (180 days after birth) at ten developmental stages were sequenced.

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Small RNA-seq analysis of single porcine blastocysts revealed that maternal estradiol-17beta exposure does not affect miRNA isoform (isomiR) expression

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