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project date 20/07/2012. The sRNA Workbench. Matthew. B. Stocks. 01. PROJECT AIMS. Based on original algorithms developed for the web based UEA sRNA Tools Moxon et. al. (2008) Easy to use
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project date20/07/2012 The sRNA Workbench Matthew. B. Stocks 01
PROJECT AIMS • Based on original algorithms developed for the web based UEA sRNA Tools • Moxon et. al. (2008) • Easy to use • Designed to be used by biologists with limited access to bioinformatics support, perhaps on a desktop computer (but is extendable to servers or high performance clusters) • Cross platform support • To allow multiple analysis techniques from within a single program through the use of an MDI (Multiple Document Interface) • Performance improvements, Speed, Memory • Still accessible through the command line • Code maintainability 02
HELPER TOOLS • Adapter Removal • Removes 3’ and/or 5’ adaptors from raw high-throughput sequence data • FASTA or FASTQ • Provides sRNA length distribution • Filter • Filters sRNA sequences from high-throughput data according to user-defined criteria • Sequence Alignment • Aligns sRNA Sequences to a Reference Genome or other long read file DEFAULT STYLES 04
miRCat • miRCat (micro RNA Categorisation) • Moxon et. al (2008) Stocks et. al. (2012) • Identify novel miRNAs in high-throughput sequence data • Identify known miRNAs found in miRBase in result set • Griffiths-Jones (2010) • View miRNA statistics • One click viewing of the predicted miRNA as they appear on the genome • One click viewing of miRNA pre-cursor secondary structure plot • Concurrency.... 06
miRCat threading CPU Core sRNA sRNA sRNA CPU Core Thread Pool: FIFO data structure (Queue) First sequence added to the list (First In) is the first item processed (First Out) sRNA Database sRNA 07
TARGET PREDICTION • Part of a parallel BBSRC tools and resources project • A high-throughput technique known as Parallel Analysis of RNA Ends (PARE) is used to sequence mRNA cleavage products on a large-scale • Sequence 5' end of uncapped mRNA (Degradome) Including transcripts targeted by sRNAs and subjected to endonucleolytic cleavage • The sRNA and degradome data can be used to identify interactions between sRNAs and their target mRNA 09
PARESnip • PAREsnip can be used to search for genome-wide interactions between all sRNAs and transcripts as well as predicting targets of small groups of miRNAs • PAREsnip outputs all potential sRNA target duplexes evidenced through the degradome • Features: • Categorisation system • Addo-Quaye et. al (2009) • Data structures based on m-way search trees and multi-threaded optimisation • ~90 mins to process 100k sequences of A. thaliana data • At least as sensitive to detecting targets as other tools designed for this type of analysis or better • Folkes et. al. (2012) NAR 10
PARESnip • Each coloured rectangle represents an interaction • The taller the rectangle, the higher the abundance • Tool tips provide additional information • T-plots can also be generated • Target information is given on the right • Navigation controls allow the user to simply click through the plots or output them to PDF for publication 11
OTHER TOOLS • miRProf (micro RNA Profiler) • Detect all known miRNAs found in miRBase within your dataset • Griffiths-Jones, (2010) • Determines normalised expression levels of known miRNA found in miRBase • Can be used to compare expression levels across multiple samples • TA-SI prediction (Trans Acting Small Interfering RNA prediction) • Predicts phased ta-siRNA in plant datasets • Chen et. al. (2007) • Requires sRNA database and genome database • View phased sRNAs as they appear on the genome • SiLoCo... • Multi sample general loci prediction • Expression profile DEFAULT STYLES 12
STATS • Since launch on 27th Jan 2011 • Total Visitors: 3,925 • Page Views: 14,650 (3.73 pages per visit) • Average time on site: 4.27 mins • Total Version 2 Downloads: 1,200 • Around 30 collated requests for the RSS feed per day 20
STATS • Visits from 57 countries/territories • UK top visitor then USA... 21
PUBLICATIONS Stocks, M. B.; Moxon, S.; Mapleson, D.; Woolfenden, H. C.; Mohorianu, I.; Folkes, L.; Schwach, F.; Dalmay, T. & Moulton, V. (2012) The UEA sRNA Workbench: A Suite of Tools for Analysing and Visualising Next Generation Sequencing microRNA and Small RNA Datasets.Bioinformatics Folkes, L.; Moxon, Simon.; Woolfenden, H.C.; Stocks, M.B.; Szittya, G.; Dalmay, T.: Moulton, V.; (2012) PAREsnip: a tool for rapid genome-wide discovery of small RNA/target interactions evidenced through degradome sequencing. Nucleic Acids Research 22
ACKNOWLEDGEMENTS Vincent Moulton Tamas Dalmay Simon Moxon Frank Schwach Irina Mohorianu Dan Mapleson Hugh Woolfenden Leighton Folkes 23