Wally

Plotting of aligned sequencing reads in BAM/CRAM format and visualization of genomic variants.

Installing Wally

The easiest way to get Wally is to download the statically linked binary from the Wally github release page. You can also build Wally from source using a recursive clone and make.

git clone --recursive https://github.com/tobiasrausch/wally.git

cd wally/

make all

Running Wally

Wally needs a sorted and indexed BAM/CRAM file and a reference genome. The output is a genomic alignment plot of the region specified on the command-line.

wally region -r chrA:35-80 -g <genome> <input.bam>

You can specify a plot name using another colon separator

wally region -r chrA:35-80:myplot -g <genome> <input.bam>

Most often you probably want to use a BED file with regions of interest and just execute wally in batch. The 4-th column specifies the plot name. For instance to plot variants from a VCF file you can use:

bcftools query -f "%CHROM\t%POS\n" <input.vcf.gz> | awk '{print $1"\t"($2-50)"\t"($2+50)"\tid"NR;}' > regions.bed

wally region -R regions.bed -g <genome> <input.bam>

Gene annotations

Simple BED files are used to provide gene or other annotations. The BED file needs to be bgzipped and indexed via tabix (tabix -p bed input.bed.gz). The required columns are chromosome, start, end and an identifier which is displayed if there is sufficient space.

wally region -b bed/gencode.hg19.bed.gz -r chr17:7573900-7574000 -g <genome> <input.bam>

The chromosome names of the genome FASTA file, BAM file and BED annotation file need to match, "chr11" and "11" are not considered identical.

Multiple BAM files

You can include multiple BAM files in a plot such as a tumor genome and a matched control in cancer genomics.

wally region -r chr17:7573900-7574000 -g <genome> <tumor.bam> <control.bam>

In fact, wally can be used to create "wallpapers" of genomes which gave rise to the tool name. For instance, the below command can be used to create a full genome view of 96 samples of a full plate of SARS-CoV-2 genomes.

wally region -y 20480 -r NC_045512.2:1-30000 -g NC_045512.2.fa Plate*.bam

Aligning multiple regions horizontally (split view)

Wally allows concatenating images of different regions horizontally using the --split option. This can be used, for instance, to zoom into a specific variant. On the command line the regions need to be separated by , without spaces. As an example, you can zoom into the N501Y variant of the alpha SARS-CoV-2 lineage using

wally region -s 3 -x 2048 -r NC_045512.2:22000-24000,NC_045512.2:23000-23100,NC_045512.2:23050-23070 -g NC_045512.2.fa <input.bam>

You can split horizontally and vertically at the same time to view, for instance, a somatic inter-chromosomal translocation.

wally region -r chrA:35-80,chrB:60-80 -g <genome> <tumor.bam> <control.bam>

If you specify the regions in a BED file using the -R option then the split parameter operates row-wise, e.g., for -s 3 row 1-3 of the BED file make up the first image, row 4-6 the second image, and so on.

Paired-end view

With -p you can switch on the paired-end view.

wally region -p -r chrA:35-80 -g <genome> <input.bam>

The paired-end coloring highlights candidate structural variants supported by read1 (R1) and read2 (R2). Below is a mapping of delly's structural variant types to wally's paired-end coloring. For inter-chromosomal translocations I assumed for this illustration that R1 maps to chromosome A and R2 maps to chromosome B.

• inversion-type paired-end, --R1--> --R2-->, INV:3to3
• inversion-type paired-end, <--R1-- <--R2--, INV:5to5
• deletion-type paired-end, --R1--> <--R2--, DEL:3to5
• duplication-type paired-end, <--R1-- --R2-->, DUP:5to3
• inter-chr paired-end, A:--R1--> B:--R2--> leads to --A--> <--B-- junction, BND:3to3
• inter-chr paired-end, A:<--R1-- B:<--R2-- leads to <--A-- --B--> junction, BND:5to5
• inter-chr paired-end, A:--R1--> B:<--R2-- leads to --A--> --B--> junction, BND:3to5
• inter-chr paired-end, A:<--R1-- B:--R2--> leads to <--A-- <--B-- junction, BND:5to3

For large and complex structural variants, wally supports split views (as explained above). For instance, for an inter-chromosomal translocation you probably want to use a 2-way horizontal split.

wally region -up -x 2048 -s 2 -r chrA:35-80,chrB:60-80 -g <genome> <tumor.bam> <control.bam>

License

Wally is distributed under the BSD 3-Clause license. Consult the accompanying LICENSE file for more details.

Credits

Wally relies heavily on the HTSlib and OpenCV. The visualization of genomic alignments was heavily inspired by IGV.