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1. Ran bwa index -a bwtsw GCF_002127325.2_HanXRQr2.0-SUNRISE_genomic.fna by hand in an interactive node (took roughly one hour)

2. Commands are in 0_assem.nf. Run this with nextflow run -bg 0_assem.nf -c teton.config. These are jobs are using: module load swset/2018.05 gcc/7.3.0 bwa/0.7.17 samtools/1.12 as specified in teton.config in this directory (bwa is version 0.7.17-r1188). Output is in /project/microbiome/data/seq/HMAX1/assem/sambam/. Gave each job 60 minutes, which was unnecessarily long, but conservative. Longest running jobs I could see were less than 20 minutes. Moved all 468 inputs files through in about 30 minutes total.

3. I removed the duplicative sam and unsorted bam files with: rm -f *.sam *[^d].bam, saving ~270 GB of space

Variant calling

• Following steps from https://github.com/zgompert/DimensionsExperiment.

• Built bcftools version 1.16 and installed in /project/evolgen/bin/.

• bcftools needed reference genome in bzip2 format, not gzip. So I now simply have an unzipped reference genome, which I have reindexed.

• Completed this step with something like: sbatch --account=evolgen --time=1-00:00 --nodes=1 --mem=8G --mail-type=END  0_call_variants.sh (this took 12 hours and 40 minutes and 552 MB of RAM; I asked for 120GB, which likely gave me the whole node and made it a bit faster)

• Filtered vcf with 1_filter_variants.sh, which contains notes on the criteria that I used (could be altered to suit).

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