HPC ALICE BLASTP

By Belmin Bajramovic @B-Bajramovic

This guide shows common BLASTP use cases using built-in BLAST options.

Preparation before running BLAST

Activate the BLAST module

module load BLAST+/2.16.0-gompi-2024a

run this to ensure the database path is correctly used

export BLASTDB=/zfsstore/databases/NCBI

To automate this for every login, add it to your bashrc

echo 'export BLASTDB=/zfsstore/databases/NCBI' >> ~/.bashrc
echo 'module load BLAST+/2.16.0-gompi-2024a' >> ~/.bashrc
source ~/.bashrc

1. Single protein vs entire nr database

Use this when you want to identify a protein or find homologs broadly.

blastp \
  -query protein.faa \
  -db /zfsstore/databases/NCBI/nr/nr \
  -evalue 1e-5 \
  -max_target_seqs 20 \
  -num_threads 8 \
  -out protein_vs_nr.tsv \
  -outfmt "6 qseqid sacc pident length qcovs evalue bitscore stitle"

2. Single protein vs a specific taxonomic group

Restrict results to a clade, order, genus, or species using NCBI taxids.
NOTE: Taxonomy filtering limits biological scope, not database size.

blastp \
  -query protein.faa \
  -db /zfsstore/databases/NCBI/nr/nr \
  -taxids 28211 \
  -evalue 1e-5 \
  -max_target_seqs 20 \
  -num_threads 8 \
  -out protein_vs_taxid.tsv \
  -outfmt "6 qseqid sacc pident length qcovs evalue bitscore stitle"

3. Searching only a specific region of the protein

Use this when only part of the protein is biologically relevant.
Residue numbering is 1-based and inclusive.

blastp \
  -query protein.faa \
  -query_loc 120-240 \
  -db /zfsstore/databases/NCBI/nr/nr \
  -evalue 1e-5 \
  -max_target_seqs 20 \
  -num_threads 8 \
  -out protein_region_120_240.tsv \
  -outfmt "6 qseqid sacc pident length qcovs evalue bitscore stitle"

4. Searching for divergent or remote homologs

Use this when close homologs are absent. This increases sensitivity at the cost of speed and specificity.

blastp \
  -query protein.faa \
  -db /zfsstore/databases/NCBI/nr/nr \
  -word_size 2 \
  -matrix BLOSUM45 \
  -evalue 1e-2 \
  -qcov_hsp_perc 30 \
  -max_target_seqs 200 \
  -num_threads 8 \
  -out divergent_homologs.tsv \
  -outfmt "6 qseqid sacc pident length qcovs evalue bitscore stitle"

5. Region-specific search for divergent homologs

Combines region restriction with relaxed similarity thresholds.

blastp \
  -query protein.faa \
  -query_loc 120-240 \
  -db /zfsstore/databases/NCBI/nr/nr \
  -word_size 2 \
  -matrix BLOSUM45 \
  -evalue 1e-2 \
  -qcov_hsp_perc 30 \
  -max_target_seqs 200 \
  -num_threads 8 \
  -out region_divergent.tsv \
  -outfmt "6 qseqid sacc pident length qcovs evalue bitscore stitle"

6. Performance and scaling guidelines

• Single protein: 1 job, 4–8 threads, <=15 minutes walltime

• Multiple proteins: use job arrays (one query per job)

• When setting cpus for your slurm job, make sure they are the same as number of threads used in blastp

• Avoid running BLAST on login nodes

Summary of key options

Use case	Key options
Broad annotation	default blastp vs nr
Taxonomy-restricted	-taxids
Region-specific	-query_loc
Divergent homologs	-word_size, -matrix, -evalue, -qcov_hsp_perc
Throughput	job arrays + -num_threads

Large query searches

To run blast for many query inputs, you should not use a single job. Instead, submit one job per query using my sbatch python script. You can download it from github and run with -h to see how to run.

git clone https://github.com/B-Bajramovic/BLAST_ALICE.git
cd BLAST_ALICE
python blast_sbatch.py -h