PacBio Software Overview

Transcript

1. FIND MEANING IN COMPLEXITY © Copyright 2013 by Pacific Biosciences of California, Inc. All rights reserved. PacBio® Software Overview
2. Learning Objectives 2 After the training, the participant will know: • The names of the PacBio software programs • Which software to use for a specific need Bioinformaticians: • Interested in understanding how the different PacBio® software programs fit within the sequencing workflow • None
3. PacBio® Software Suite 3 • Fully automated analysis from run setup, with the option to manually run later • Efficient integration with LIMS and third-party analysis tools • User-friendly UI design for advanced informatics researchers, as well as biologists and clinicians • Industry-standard output formats: FASTA, FASTQ, SAM/BAM, VCF • Open source through our developers’ network ®
4. High Level Software Workflow 4 Laboratory Information Management System API Layer Primary Analysis Secondary Analysis Tertiary Analysis Sample Preparation Design Run Load Instrument RS Touch and RS Remote SMRT® Portal and SMRT® View Run & Monitor Review Run
5. RS Touch and RS Remote 5 Primary Analysis Design Run Load Instrument Run & Monitor Review Run RS Touch RS Remote RS Remote Secondary Analysis Tertiary Analysis Sample Preparation RS Touch RS Remote
6. RS Remote – Designing a Run 6
7. RS Remote – Designing a Run 7 • Design runs remotely • Can assign multiple SMRT® Cells per well with different movie times
8. RS Touch – Loading a Run 8
9. RS Touch – Monitoring a Run 9 • Monitor at the instrument or remotely • View real-time base incorporations • Displays remaining run time • Shows SMRT® Cell status, from SMRT Cell prep to Quality Values
10. PacBio® High Resolution Genetic Analyzer 10
11. Movie Trace Pulse Raw Base Calls CC Base Calls SMRT® Cell Movie2Trace Trace2Pulse Pulse2Base Base2 CircularConsensus 2° Analysis Primary Analysis Pipeline 11 GCAACGATCA…GCAACGATCA…GCAACGATCA…GCAACGATCA…GCAA ACGATCA Pulse Features • Pulse Height • Pulse Width • Interpulse Distance t
12. High Performance Computing Analysis Setup 12 Data Repository Customer LAN PacBio® High Resolution Genetic Analyzer Base calls + QVs Primary Analysis: Signal processing, base calling, quality assessment Secondary Analysis: Alignment/Assembly
13. RS Insight – Remote Monitoring • Proactive monitoring by PacBio Tech Support if RS Insight access is enabled – Sequencing QC metrics are retained on the Blade Center 13 Service Management Tech Support Field Service R & D Customer- Controlled Policy Server Internet PacBio® RS PacBio Remote Engineering Team
14. SMRT® Analysis – Secondary Analysis 14 Secondary Analysis SMRT Portal Alignment/Assembly SMRT View Visualization Primary Analysis Design Run Load Instrument Run & Monitor Review Run Tertiary Analysis Sample Preparation
15. SMRT® Portal 15 • Web-based interface • Accessible from any computer • Automated secondary analysis
16. SMRT® Portal – Job Selection 16 • Prior jobs are searchable • Can select one or more jobs
17. SMRT® Portal – Report Viewing 17 Full complement of reports automatically generated Quality Values Readlength distributions Post-secondary accuracy distributions Automatic generation of industry- standard data formats FASTA/FASTQ SRA SAM/BAM …others Integrated genome browser Direct connection to SMRT® View
18. SMRT® View – Genome Browser 18
19. SMRT® View Base Modification Details 19 Kinetogram: IPD ratio by base position and strand Modification density by mod, variants, and separate tracks per motif
20. Application-Specific Analysis Software De Novo Assembly HGAP: Generate high quality assemblies from PacBio® long reads alone ALLORA: Assemble pure PacBio long reads, then polish with Quiver AHA: Fill gaps and join existing scaffolds with PacBio long reads Celera® Assembler: Combine PacBio long reads with short reads from other technologies SMRT® View: QC assemblies 20 Targeted Sequencing BLASR: Map reads against a reference Quiver: Call haploid SNPs and indels with 99.999% accuracy GATK: Identify haploid and diploid SNPs using the Broad’s Unified Genotyper GMAP: Align full-length cDNA transcripts against genomic DNA to discover splicing SMRT View: Browse coverage, variants and annotations DNA Base Modifications Available as an extension of standard resequencing, or using a case/control comparison Modification detection: Find specific modified sites in unamplified genomes Bacterial methylomes: Discover recognition motifs for adenine and cytosine methylation SMRT View: Visualize modified sites and sequence contexts
21. DevNet 21
22. Summary of Key Points • Suite of data management and analysis software tools - Supports granularity, scalability, and full functionality of the PacBio® RS instrument • Integrated software solution from beginning to end - Fully automated analysis, with the option for manual set-up • Accessible and usable - User-friendly UI design • Industry-standard output formats - FASTA, FASTQ, SAM/BAM, and so on. • Open and extensible platform - Developer’s Network 22
23. Where to Find More Information These documents are all available on DevNet: – SMRT Portal Help – SMRT View Help – SMRT Pipe Reference Guide (v1.4) – SMRT Analysis Software Installation (v1.4) – Software Getting Started Guide 23