This document discusses assembly tools and visualization software. It describes assemblers like ABySS and SOAPdenovo that use de Bruijn graphs to assemble genomes from short reads. It also discusses tools for visualizing assemblies, like Tablet and ABySS-Explorer. Finally, it covers read mapping with SAM/BAM formats and tools like BWA, and visualization of mappings with Artemis and IGV.

1. Assembly tools and Visualisation
Matthias Haimel
EBI is an Outstation of the European Molecular Biology Laboratory.

2. Overview
• Assemblers
• ABySS
• SOAPdenovo
• Visualisation
• Tablet
• AbySS-Explorer
• Read mapping
• Sam / Bam
• Visualisation
• Artemis
• IGV - Integrative Genomics Viewer
2 25.04.11 Assemblers

3. ABySS Assembly By Short Sequences
• Genome Sciences Centre, Vancouver
• http://www.bcgsc.ca/platform/bioinfo/software/abyss
• Open source, BCCA Licence
• de Bruijn graph
• Trimming (tip clipping), bubble popping
• Use paired-end information: resolve ambiguities between contigs
• parallel (use cluster)
• Files
• Fasta / Fastq
• Sam/Bam
• colour-space
3 25.04.11 Assemblers

4. ABySS
• ABYSS (singe end)
• e.g. ABYSS -k27 single.fastq -o contigs.fa
• abyss-pe (paired end)
• e.g. abyss-pe k=27 n=10 in='read_1.fastq read_2.fastq' name=ecli
• Multiple libraries
• ... lib=’read1 read2’ read1=’read1_1.fa read1_2.fa’ read2=’read2_1.fa read1_2.fa’
4 25.04.11 Assemblers

5. SOAPdenovo
• Beijing Genomics Institute (BGI), China
• http://soap.genomics.org.cn/soapdenovo.html
• Panda genome
• Source available
• de Bruijn graph
• pre-set Kmer frequency threshold
• Bubble removing
• Build scaffold
• mapping reads to contigs
• gap filling
5 25.04.11 Assemblers

6. SOAPdenovo
• Full run
• e.g. SOAPdenovo all -s read.config -K 27 -o contigs.fa
• Run sub steps
• pregraph = velveth
• contig = velvetg
• map map reads to contigs
• scaff scaffolding
• Configuration
• Config file input instead of read files
• Specify rank, usage (assembly/scaffolding), insert size
6 25.04.11 Assemblers

7. Visualisation http://bioinf.scri.ac.uk/tablet/
• Tablet
• Lightweight
• Easy to use
• Formats
• ACE
• AFG
• BAM
• BANK (AMOS)
7 25.04.11 Assemblers

8. Visualisation - Velvet
• Tablet
• Velvetg ... -amos_file yes
• GraphViz
• Transform velvet graph into GraphViz format
• Contributed by Paul Harrison
• <velvet>/contrib/layout/
• Velvet -> .dot file (Python script)
• .dot -> png (graphviz)
8 25.04.11 Assemblers

9. Visualisation http://www.bcgsc.ca/platform/bioinfo/software/abyss-explorer
• ABySS-Explorer
• Visualizes ABySS assemblies
• Interactive graph structure
• Filter contigs
9 25.04.11 Assemblers

10. Assembler - Practical
• Assemblers
• ABySS
• SOAPdenovo
• Visualisation
• Tablet
• ABySS-Explorer
10 25.04.11 Assemblers

11. Read mapping http://samtools.sourceforge.net/SAM1.pdf
• SAM / BAM
• Sequence Alignment / Map format (SAM)
• Binary form of SAM (BAM)
• generic format
• Flexible and simple
• Compact (BAM)
• Allow indexing
• Load regions
• Support streaming
11 25.04.11 Assemblers

12. SAM
• Header
• File format version information
• Sequence dictionary (name/length/..)
• Read group (platform/library/...)
• Program info
• Body
• Alignment information
12 25.04.11 Assemblers

13. SAM Header
• '@' followed by record type (two characters)
@HD VN:1.0
@SQ SN:chr20 LN:62435964
@RG ID:L1 PU:SC_1_10 LB:SC_1 SM:NA12891
@RG ID:L2 PU:SC_2_12 LB:SC_2 SM:NA12891
13 25.04.11 Assemblers

14. SAM Alignment
• Tab delimited lines
14 25.04.11 Assemblers

15. SAM Alignment
• Tab delimited lines
Read_28833_29006_6945 99 chr20 28833 20 10M1D25M = 28993 195
AGCT... <<<<... NM:i:1 RG:Z:L1
read_28701_28881_323b 147 chr20 28834 30 35M = 28701 -168
ACCT... <<7;:... MF:i:18 RG:Z:L2
15 25.04.11 Assemblers

16. Tools
• Mapping Reads
• BWA
• Bowtie
• SSAHA2
• Manipulate SAM/BAM
• SAM Tools package
• Picard
16 25.04.11 Assemblers

17. BWA
• Burrows-Wheeler Alignment Tool
• Map (singe/paired-end/long) reads to a sequence
• Index database
• bwa index -a bwtsw database.fasta
• Align reads
• bwa aln database.fasta short_read.fastq > aln_sa.sai
• Generate alignments
• bwa sampe database.fasta aln_sa1.sai aln_sa2.sai read1.fq read2.fq > aln.sam
• Long reads
• bwa bwasw database.fasta long_read.fastq > aln.sam
17 25.04.11 Assemblers

18. SAM tools
• Utilities for SAM format
• samtools <command> ...
• Commands:
• view: SAM <-> BAM
• sort: sort BAM file
• index: build BAM file index
• merge: merges x BAM files
• pileup: alignment in the pileup format
• tview: integrated Text alignment viewer
18 25.04.11 Assemblers

19. Visualisation Integrative Genomics Viewer
http://www.broadinstitute.org/igv/
• IGV
• Good integration
• Formats
• DAS
• BAM
• GFF
• ...
• Tools
• Run scripts
• Export region
• ...
19 25.04.11 Assemblers

20. Visualisation
http://www.sanger.ac.uk/resources/software/artemis/
• Artemis
• Sequence Viewer
• Annotation tool
• Formats
• EMBL
• GENBANK
• GFF
• FASTA
• BAM
20 25.04.11 Assemblers

21. Mapping - Practical
• Mapping reads + prepare for visalization
• BWA
• samtools
• Visualisation
• IGV
21 25.04.11 Assemblers