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# Installation of software for trio whole exome project
## Aspera
Downloaded Aspera Connect version 3.7.4.147727 from https://downloads.asperasoft.com and extracted to /home/u035/project/software.
Version 1.2.3 with some bugfixes from the dev branch as of 26 August 2020.
Start with installing the base software, and add datatargets.
This will take a long time, and may require multiple runs if it fails on a step. It will resume if needed. Run on a screen session and log each attempt. It's important to set the limit on the number of concurrently open files to as high as possible (4096 on ultra).
```
cd /home/u035/project/software/install
wget https://raw.github.com/bcbio/bcbio-nextgen/master/scripts/bcbio_nextgen_install.py
DATE=`date +%Y%m%d%H%M`
python bcbio_nextgen_install.py /home/u035/project/software/bcbio \
--cores 32 &> bcbio_install_base_${DATE}.log
Fix an issue with bcbio & vt/samtools/htslib. See https://github.com/bcbio/bcbio-nextgen/issues/3327 and https://github.com/bcbio/bcbio-nextgen/issues/3328.
DATE=`date +%Y%m%d%H%M`
/home/u035/project/software/bcbio/tools/bin/bcbio_nextgen.py upgrade -u development --tools &> bcbio_install_upgrade_tools_${DATE}.log
Install datatarget vep
DATE=`date +%Y%m%d%H%M`
/home/u035/project/software/bcbio/tools/bin/bcbio_nextgen.py upgrade -u skip --datatarget vep &> bcbio_install_datatarget_vep_${DATE}.log
We already had gnomAD 3.0 compiled and downloaded from another bcbio installation, so this gets copied to /home/u035/project/software/bcbio/genomes/Hsapiens/hg38/variation. However, if needed, re-generate it like this. It will take about 6 days.
```
DATE=`date +%Y%m%d%H%M`
/home/u035/project/software/bcbio/tools/bin/bcbio_nextgen.py upgrade -u skip --datatarget gnomad &> bcbio_install_datatarget_gnomad_${DATE}.log
```
Increase JVM memory for GATK in galaxy/bcbio_system.yaml
```
gatk:
jvm_opts: ["-Xms500m", "-Xmx5g"]
```
### Patch Ensembl VEP 100.4
See https://github.com/Ensembl/ensembl-variation/pull/621/files
Edit /home/u035/project/software/bcbio/anaconda/share/ensembl-vep-100.4-0/Bio/EnsEMBL/Variation/BaseAnnotation.pm accordingly.
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### Verifybamid custom panel for exomes
```
source /home/u035/project/scripts/trio_whole_exome_config.sh
mkdir /home/u035/project/software/install/1000G_phase3_hg38
cd /home/u035/project/software/install/1000G_phase3_hg38
# download the 1000 Genomes autosomes + X site VCFs
for ((i = 1; i <= 22; i = i + 1))
do
wget ftp://ftp.1000genomes.ebi.ac.uk/vol1/ftp/data_collections/1000_genomes_project/release/20190312_biallelic_SNV_and_INDEL/ALL.chr${i}.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.vcf.gz;
wget ftp://ftp.1000genomes.ebi.ac.uk/vol1/ftp/data_collections/1000_genomes_project/release/20190312_biallelic_SNV_and_INDEL/ALL.chr${i}.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.vcf.gz.tbi
done
wget ftp://ftp.1000genomes.ebi.ac.uk/vol1/ftp/data_collections/1000_genomes_project/release/20190312_biallelic_SNV_and_INDEL/ALL.chrX.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.vcf.gz
wget ftp://ftp.1000genomes.ebi.ac.uk/vol1/ftp/data_collections/1000_genomes_project/release/20190312_biallelic_SNV_and_INDEL/ALL.chrX.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.vcf.gz.tbi
cd ..
# create bare to prefixed chromosome map
for ((i = 1; i <= 22; i = i + 1))
do
echo $i "chr"$i >> chr_prefix_map.txt
done
echo chrX >> chr_prefix_map.txt
# use the TWIST kit to subset the variants and add the chr prefix at the same time
for file in 1000G_phase3_hg38/*vcf.gz
do
bname=`basename $file`
bcftools view -R /home/u035/project/resources/Twist_Exome_Target_hg38.bed -m2 -M2 -v snps -i 'AF >= 0.01' $file | bcftools annotate --rename-chrs chr_prefix_map.txt | bgzip -c > ${bname%.vcf.gz}.biallelic.snps.m\
inAF0.01.vcf.gz
tabix ${bname%.vcf.gz}.biallelic.snps.minAF0.01.vcf.gz
done
# concatenate all the files in the correct order
bcftools concat -o ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.chr.biallelic.snps.minAF0.01.vcf.gz -O z \
ALL.chr[1-9].shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.chr.biallelic.snps.minAF0.01.vcf.gz \
ALL.chr[12][0-9].shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.chr.biallelic.snps.minAF0.01.vcf.gz \
ALL.chrX.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.chr.biallelic.snps.minAF0.01.vcf.gz
tabix ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.chr.biallelic.snps.minAF0.01.vcf.gz
# use VerifyBamID to create the new panel
/home/u035/project/software/bcbio/anaconda/share/verifybamid2-1.0.6-0/VerifyBamID \
--RefVCF ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.chr.biallelic.snps.minAF0.01.vcf.gz
--Reference bcbio-1.1.5/genomes/Hsapiens/hg38/seq/hg38.fa
# rename the files to the correct format
mv ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.chr.biallelic.snps.minAF0.01.vcf.gz.bed 1000g.phase3.100k.b38.vcf.gz.dat.bed
mv ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.chr.biallelic.snps.minAF0.01.vcf.gz.mu 1000g.phase3.100k.b38.vcf.gz.dat.mu
mv ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.chr.biallelic.snps.minAF0.01.vcf.gz.PC 1000g.phase3.100k.b38.vcf.gz.dat.V
mv ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.chr.biallelic.snps.minAF0.01.vcf.gz.UD 1000g.phase3.100k.b38.vcf.gz.dat.UD
# move them into the correct location, backing up the original resource folder
cd /home/u035/project/software/bcbio/anaconda/share/verifybamid2-1.0.6-0
mv resource resource.bak
mkdir resource
mv /home/u035/project/software/install/1000G_phase3_hg38/1000g.phase3.100k.b38* resource/
```
## Python modules
### VASE
cd /home/u035/project/software
./bcbio/anaconda/bin/pip3 install git+git://github.com/david-a-parry/vase.git#egg=project[BGZIP,REPORTER,MYGENE]
XlsxWriter 1.3.3 was installed 28 August 2020.
cd /home/u035/project/software
./bcbio/anaconda/bin/pip3 install XlsxWriter
## GATK 3.8
```
cd /home/u035/project/software/install
wget https://storage.googleapis.com/gatk-software/package-archive/gatk/GenomeAnalysisTK-3.8-0-ge9d806836.tar.bz2
bzip2 -d GenomeAnalysisTK-3.8-0-ge9d806836.tar.bz2
tar -xf GenomeAnalysisTK-3.8-0-ge9d806836.tar
mv GenomeAnalysisTK-3.8-0-ge9d806836 ../GenomeAnalysisTK-3.8
```