title: iMir: an integrated pipeline for high-throughput analysis of small non-coding RNA data obtained by smallRNA-Seq.
abstract: BACKGROUND
  Qualitative and quantitative analysis of small non-coding RNAs by next generation sequencing (smallRNA-Seq) represents a novel technology increasingly used to investigate with high sensitivity and specificity RNA population comprising microRNAs and other regulatory small transcripts. Analysis of smallRNA-Seq data to gather biologically relevant information, i.e. detection and differential expression analysis of known and novel non-coding RNAs, target prediction, etc., requires implementation of multiple statistical and bioinformatics tools from different sources, each focusing on a specific step of the analysis pipeline. As a consequence, the analytical workflow is slowed down by the need for continuous interventions by the operator, a critical factor when large numbers of datasets need to be analyzed at once.
  
  
  RESULTS
  We designed a novel modular pipeline (iMir) for comprehensive analysis of smallRNA-Seq data, comprising specific tools for adapter trimming, quality filtering, differential expression analysis, biological target prediction and other useful options by integrating multiple open source modules and resources in an automated workflow. As statistics is crucial in deep-sequencing data analysis, we devised and integrated in iMir tools based on different statistical approaches to allow the operator to analyze data rigorously. The pipeline created here proved to be efficient and time-saving than currently available methods and, in addition, flexible enough to allow the user to select the preferred combination of analytical steps. We present here the results obtained by applying this pipeline to analyze simultaneously 6 smallRNA-Seq datasets from either exponentially growing or growth-arrested human breast cancer MCF-7 cells, that led to the rapid and accurate identification, quantitation and differential expression analysis of ~450 miRNAs, including several novel miRNAs and isomiRs, as well as identification of the putative mRNA targets of differentially expressed miRNAs. In addition, iMir allowed also the identification of ~70 piRNAs (piwi-interacting RNAs), some of which differentially expressed in proliferating vs growth arrested cells.
  
  
  CONCLUSIONS
  The integrated data analysis pipeline described here is based on a reliable, flexible and fully automated workflow, useful to rapidly and efficiently analyze high-throughput smallRNA-Seq data, such as those produced by the most recent high-performance next generation sequencers. iMir is available at http://www.labmedmolge.unisa.it/inglese/research/imir.
authors: Giorgio
authors: G
authors: Giurato
authors: Maria Rosaria
authors: MR
authors: De Filippo
authors: Antonio
authors: A
authors: Rinaldi
authors: Adnan
authors: A
authors: Hashim
authors: Giovanni
authors: G
authors: Nassa
authors: Maria
authors: M
authors: Ravo
authors: Francesca
authors: F
authors: Rizzo
authors: Roberta
authors: R
authors: Tarallo
authors: Laboratory of Molecular Medicine and Genomics, Department of Medicine and Surgery, University of Salerno, via Allende, 1, Salerno, Baronissi, Italy. aweisz@unisa.it.
authors: Alessandro
authors: A
authors: Weisz
mesh_terms: D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D018390:Gene Targeting / Q000379:methods; D005815:Genetic Code / Q000235:genetics; D055106:Genome-Wide Association Study / Q000379:methods; D023281:Genomics / Q000379:methods*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D061986:MCF-7 Cells; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D011237:Predictive Value of Tests; D058727:RNA, Small Untranslated / Q000235:genetics* / Q000378:metabolism; D017423:Sequence Analysis, RNA / Q000379:methods*; D012984:Software; D014584:User-Computer Interface
chemical_list: D035683:MicroRNAs; D058727:RNA, Small Untranslated