How-to: align DNA-seq data
Aligning with BWA
References: https://www.broadinstitute.org/files/shared/mpg/nextgen2010/nextgen_li.pdf https://genomics.sschmeier.com/ngs-mapping/#mapping-statistics
Get BWA
http://sourceforge.net/projects/bio-bwa/files/ http://bio-bwa.sourceforge.net/bwa.shtml
bunzip2 bwa-0.X.X.tar.bz2
tar xvf bwa-0.X.X.tar
cd bwa-0.X.X
make
export PATH=$PATH:/path/to/bwa-0.X.X
source ~/.bashrc
Download data
Reference genome
wget ftp://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_31/GRCh38.p12.genome.fa.gz
Index genome
bwa index -a bwtsw GRCh38.genome.fa
Mapping
Mapping Short reads
Paired-end
bwa aln -t 4 GRCh38bwaidx reads1.fastq.gz > reads1.bwa
bwa aln -t 4 GRCh38bwaidx reads2.fastq.gz > reads2.bwa
bwa sampe GRCh38bwaidx reads1.bwa reads2.bwa reads1.fastq.gz reads2.fastq.gz > aln.sam
Single-end
bwa aln -t 4 GRCh38bwaidx reads.fastq.gz > reads.bwa
bwa samse GRCh38bwaidx reads.bwa reads.fastq.gz > aln.sam
. Mapping long reads
bwa bwasw GRCh38bwaidx long-reads.fastq.gz > aln-long.sam
. Mapping with BWA-MEM
bwa index GRCh38.genome.fa
Paired-end
bwa mem GRCh38.genome.fa reads1.fastq.gz reads2.fastq.gz > aln-pe.sam
Single-end
bwa mem GRCh38.genome.fa reads.fq.gz > aln-se.sam
Processing
Fix mates and compress
samtools sort -n -O sam aln.sam | samtools fixmate -m -O bam - aln.fixmate.bam
Sorting
samtools sort -O bam -o aln.sorted.bam aln.fixmate.bam
Remove duplicates
samtools markdup -r -S aln.sorted.bam aln.sorted.dedup.bam
