# Ultra2 - Installation of software for trio whole exome project

## Aspera

Downloaded Aspera Connect version 3.9.6.1467 installer script from https://downloads.asperasoft.com to /home/u035/project/software/install and run it. This installs the software in ~/.aspera, so it needs to be moved to the shared folder.

```
bash ibm-aspera-cli-3.9.6.1467.159c5b1-linux-64-release.sh
mv ~/.aspera ../aspera
```

## bcbio

Version 1.2.8 (14 April 2021).

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).

```
cd /home/u035/u035/shared/software/install
mkdir bcbio_install_logs

wget https://raw.github.com/bcbio/bcbio-nextgen/master/scripts/bcbio_nextgen_install.py

ulimit -n 4096

DATE=`date +%Y%m%d%H%M`
python3 bcbio_nextgen_install.py /home/u035/u035/shared/software/bcbio \
  --tooldir /home/u035/u035/shared/software/bcbio/tools \
  --genomes hg38 --aligners bwa \
  --cores 128 &> bcbio_install_logs/bcbio_install_base_${DATE}.log
```

Install datatarget vep

```
DATE=`date +%Y%m%d%H%M`
/home/u035/u035/shared/software/bcbio/tools/bin/bcbio_nextgen.py upgrade -u skip --datatarget vep &> bcbio_install_logs/bcbio_install_datatarget_vep_${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/u035/shared/software/bcbio/anaconda/share/ensembl-vep-100.4-0/Bio/EnsEMBL/Variation/BaseAnnotation.pm accordingly.

### Verifybamid custom panel for exomes

```
mkdir /home/u035/u035/shared/software/install/1000G_phase3_hg38
cd /home/u035/u035/shared/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

# 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 X chrX >> chr_prefix_map.txt

# add bcbio tools to path
PATH=/home/u035/u035/shared/software/bcbio/tools/bin:/home/u035/u035/shared/software/bcbio/anaconda/share/verifybamid2-1.0.6-0:$PATH

# use the TWIST kit to subset the variants and add the chr prefix at the same time
sed -e 's/chr//' ../../../resources/Twist_Exome_Target_hg38.bed > targets.bed
for file in *phased.vcf.gz
do
  bname=`basename $file`
  bcftools view -R targets.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.minAF0.01.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.biallelic.snps.minAF0.01.vcf.gz -O z \
  ALL.chr[1-9].shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.biallelic.snps.minAF0.01.vcf.gz \
  ALL.chr[12][0-9].shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.biallelic.snps.minAF0.01.vcf.gz \
  ALL.chrX.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.biallelic.snps.minAF0.01.vcf.gz
tabix ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.biallelic.snps.minAF0.01.vcf.gz

# use VerifyBamID to create the new panel
VerifyBamID \
  --RefVCF ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.biallelic.snps.minAF0.01.vcf.gz \
  --Reference ../../bcbio/genomes/Hsapiens/hg38/seq/hg38.fa

# rename the files to the correct format
mv ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.biallelic.snps.minAF0.01.vcf.gz.bed 1000g.phase3.100k.b38.vcf.gz.dat.bed
mv ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.biallelic.snps.minAF0.01.vcf.gz.mu 1000g.phase3.100k.b38.vcf.gz.dat.mu
mv ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.biallelic.snps.minAF0.01.vcf.gz.PC 1000g.phase3.100k.b38.vcf.gz.dat.V
mv ALL.shapeit2_integrated_snvindels_v2a_27022019.GRCh38.phased.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/u035/shared/software/bcbio/anaconda/share/verifybamid2-1.0.6-0
mv resource resource.bak
mkdir resource
mv /home/u035/u035/shared/software/install/1000G_phase3_hg38/1000g.phase3.100k.b38* resource/

# clean up intermediate files
cd /home/u035/u035/shared/software/install
rm -r 1000G_phase3_hg38
```

## Python modules

### VASE

VASE v0.4.2 was installed 18 August 2021.

```
cd /home/u035/u035/shared/software
./bcbio/anaconda/bin/pip3 install git+git://github.com/david-a-parry/vase.git#egg=vase[BGZIP,REPORTER,MYGENE]
```

### XlsxWriter

XlsxWriter 3.0.1 was installed 18 August 2021.

```
cd /home/u035/u035/shared/software
./bcbio/anaconda/bin/pip3 install XlsxWriter
```

## GATK 3.8

```
cd /home/u035/u035/shared/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
rm GenomeAnalysisTK-3.8-0-ge9d806836.tar
```

## RTG tools

```
cd /home/u035/u035/shared/software
wget https://github.com/RealTimeGenomics/rtg-tools/releases/download/3.11/rtg-tools-3.11-linux-x64.zip
unzip rtg-tools-3.11-linux-x64.zip
rm rtg-tools-3.11-linux-x64.zip
```

## IGV

```
cd /home/u035/u035/shared/software
wget https://data.broadinstitute.org/igv/projects/downloads/2.8/IGV_Linux_2.8.9.zip
unzip IGV_Linux_2.8.9.zip
rm IGV_Linux_2.8.9.zip
```

## Emacs

```
cd /home/u035/u035/shared/software
./bcbio/anaconda/bin/conda install emacs
```