Using Galaxy for Molecular Assay Design

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Presentation transcript:

Using Galaxy for Molecular Assay Design 2012 Galaxy Community Conference J Ireland, Principal Bioinformatics Scientist

Galaxy at 5AM Solutions 5AM: Custom software and bioinformatics services company Galaxy converts for one year http://info.5amsolutions.com/blog/bid/86705/DIY-Bioinformatics- A-Whole-New-Galaxy Galaxy in action Internal bioinformatics resource Client 1: data source integration Client 2: custom workflows for assay design Client 3 (in progress): deploy Galaxy for assay design

What is Molecular Assay Design (MAD)? In-silico process to identify oligos (probes, primers) to detect and/or quantify nucleic acid targets Balance Target Coverage: oligo location, expected performance Cost: $$$, oligo real-estate, oligo interactions Technologies PCR, TaqMan, molecular inversion probes, LNA, oligo arrays, … Applications Targeted sequencing, genotyping, gene expression, miRNA, …

The MAD Workflow Target Selection Assay Design Identify locations on genome/transcriptome of interest while avoiding trouble regions Assay Design Select sets of primers and/or probes against targets Assay Prioritization/Filter Filter, prioritize and annotate assay designs Ensemble Selection Select final combination(s) of assays

Cell-Free DNA Testing – A MAD Example Client developing a multiplexed PCR assay for detection of trisomies in cfDNA Requires sensitive assay to detect fetal cfDNA in high background of maternal cfDNA Short assay lengths needed because cfDNA highly fragmented Genetic variation (SNPs, CNV) can confound detection Fan, et al. Clin. Chem, 2010

Targets for cfDNA Testing RefSeq Exons Segmental Duplications dbSNP

An Aside: Galaxy Powered MAD Analysis

Assay Design

Assay Prioritization and Filtering

Assay Ensembles Combine multiplexed pools Select “optimal” subset Avoid misamplification Avoid oligo interactions Select “optimal” subset Coverage Expected performance Best multiplex Programs to help MultiPLX Pool 1 Pool 2

Looking Back No more Perl soup Data sets, parameters, steps preserved Reproducible design Sharable with client

Looking Ahead Tool Shed submission Extend workflows to ensemble selection Automate report generation Sweave for Galaxy?

Thanks! Andy Evans Will FitzHugh The Galaxy Community!