Taxonomic Assignments – C.bairdi 6129-403-26-Q7 NanoPore Reads Using DIAMOND BLASTx on Mox and MEGAN6 daa2rma on emu

After noticing that the initial MEGAN6 taxonomic assignments for our combined C.bairdi NanoPore data from 20200917 revealed a high number of bases assigned to E.canceri and Aquifex sp., I decided to explore the taxonomic breakdown of just the individual samples to see which of the samples was contributing to these taxonomic assignments most.

Ran the Hematodinium-infected (6129_403_26_Q7) hemolymph NanoPore sequencing data through DIAMOND BLASTx (on Mox) and subsequent output conversion to the MEGAN6 RMA6 format (on swoose due to program Java X11 requirement which is not functional on Mox) to obtain taxonomic assignments.

SBATCH script (GitHub):

#!/bin/bash ## Job Name #SBATCH --job-name=cbai_blastx_DIAMOND_nanopore_6129_403_26_Q7 ## Allocation Definition #SBATCH --account=coenv #SBATCH --partition=coenv ## Resources ## Nodes #SBATCH --nodes=1 ## Walltime (days-hours:minutes:seconds format) #SBATCH --time=10-00:00:00 ## Memory per node #SBATCH --mem=200G ##turn on e-mail notification #SBATCH --mail-type=ALL #SBATCH --mail-user=samwhite ## Specify the working directory for this job #SBATCH --chdir=/gscratch/scrubbed/samwhite/outputs/20200928_cbai_diamond_blastx_nanopore_6129_403_26_Q7 # Script to run DIAMOND BLASTx on 6129_403_26 quality filtered (Q7) C.bairdi NanoPore reads # from 20200928 using the --long-reads option # for subsequent import into MEGAN6 to evaluate reads taxonomically. ################################################################################### # These variables need to be set by user # Input FastQ file fastq=/gscratch/srlab/sam/data/C_bairdi/DNAseq/20200928_cbai_nanopore_6129_403_26_quality-7.fastq # DIAMOND Output filename prefix prefix=20200928_cbai_nanopore_6129_403_26_Q7 # Set number of CPUs to use threads=28 # Program paths diamond=/gscratch/srlab/programs/diamond-2.0.4/diamond # DIAMOND NCBI nr database with taxonomy dumps dmnd_db=/gscratch/srlab/blastdbs/ncbi-nr-20190925/nr.dmnd ################################################################################### # Exit script if any command fails set -e # Load Python Mox module for Python module availability module load intel-python3_2017 # SegFault fix? export THREADS_DAEMON_MODEL=1 # Inititalize arrays programs_array=() # Programs array programs_array=("${diamond}") md5sum "${fastq}" > fastq_checksums.md5 # Run DIAMOND with blastx # Output format 6 produces a standard BLAST tab-delimited file # Run DIAMOND with blastx # Output format 100 produces a DAA binary file for use with MEGAN ${diamond} blastx \ --long-reads \ --db ${dmnd_db} \ --query "${fastq}" \ --out "${prefix}".blastx.daa \ --outfmt 100 \ --top 5 \ --block-size 8.0 \ --index-chunks 1 \ --threads ${threads} # Capture program options for program in "${!programs_array[@]}" do { echo "Program options for ${programs_array[program]}: " echo "" ${programs_array[program]} help echo "" echo "" echo "