1. Prioritizing and selecting likely novel miRNAs from NGS data
- Author
-
Benjamin Meder, Eckart Meese, Klemens Ruprecht, Christina Backes, Patrick Roth, Jan Haas, Martin Hart, Nicole Ludwig, Valentina Galata, Mustafa Kahraman, Friedrich A. Grässer, Andreas Keller, Britta Vogel, Jennifer Menegatti, Petra Leidinger, Thomas Grossmann, University of Zurich, and Keller, A
- Subjects
0301 basic medicine ,Sequence analysis ,Molecular Sequence Data ,610 Medicine & health ,Computational biology ,Biology ,Real-Time Polymerase Chain Reaction ,600 Technik, Medizin, angewandte Wissenschaften::610 Medizin und Gesundheit ,MiRBase ,03 medical and health sciences ,1311 Genetics ,microRNA ,Genetics ,False positive paradox ,Humans ,Base sequence ,Blood Cells ,Base Sequence ,Sequence Analysis, RNA ,Gene Expression Profiling ,Computational Biology ,High-Throughput Nucleotide Sequencing ,Blotting, Northern ,10040 Clinic for Neurology ,Gene expression profiling ,MicroRNAs ,030104 developmental biology ,Reference database ,Methods Online ,Transcriptome ,Algorithms ,Software - Abstract
Small non-coding RNAs play a key role in many physiological and pathological processes. Since 2004, miRNA sequences have been catalogued in miRBase, which is currently in its 21st version. We investigated sequence and structural features of miRNAs annotated in the miRBase and compared them between different versions of this reference database. We have identified that the two most recent releases (v20 and v21) are influenced by next-generation sequencing based miRNA predictions and show significant deviation from miRNAs discovered prior to the high-throughput profiling period. From the analysis of miRBase, we derived a set of key characteristics to predict new miRNAs and applied the implemented algorithm to evaluate novel blood-borne miRNA candidates. We carried out 705 individual whole miRNA sequencings of blood cells and collected a total of 9.7 billion reads. Using miRDeep2 we initially predicted 1452 potentially novel miRNAs. After excluding false positives, 518 candidates remained. These novel candidates were ranked according to their distance to the features in the early miRBase versions allowing for an easier selection of a subset of putative miRNAs for validation. Selected candidates were successfully validated by qRT-PCR and northern blotting. In addition, we implemented a web-server for ranking potential miRNA candidates, which is available at: www.ccb.uni-saarland.de/novomirank.
- Published
- 2016