How do I perform a local BLAST search against a custom protein database in Python?

Question

I have a set of protein sequences in a FASTA file and I want to run a local BLAST search against a custom database I built. I'm using Biopython.

Here's what I have so far:

```python
from Bio.Blast.Applications import NcbiblastpCommandline

blastp_cline = NcbiblastpCommandline(
    query='my_sequences.fasta',
    db='my_custom_db',
    evalue=0.001,
    outfmt=5,
    out='results.xml'
)
stdout, stderr = blastp_cline()
```

I get the error: `BLAST Database error: No alias or index file found for protein database`. What am I missing?

Admin · Accepted Answer

You need to first create the BLAST database using `makeblastdb` before you can query it. Here's the full workflow:

```python
from Bio.Blast.Applications import NcbimakeblastdbCommandline, NcbiblastpCommandline

# Step 1: Create the database
makeblastdb_cline = NcbimakeblastdbCommandline(
    input_file='my_protein_sequences.fasta',
    dbtype='prot',
    out='my_custom_db',
    title='My Custom Protein DB'
)
stdout, stderr = makeblastdb_cline()

# Step 2: Run BLAST
blastp_cline = NcbiblastpCommandline(
    query='my_sequences.fasta',
    db='my_custom_db',
    evalue=0.001,
    outfmt=5,
    out='results.xml'
)
stdout, stderr = blastp_cline()
```

Make sure BLAST+ is installed and accessible from your PATH. You can verify with `which blastp` on Linux/Mac.

Admin · Answer

Also worth noting: if you're running this on a large database, consider using Diamond BLAST instead — it's orders of magnitude faster for large-scale searches:

```bash
diamond makedb --in my_proteins.fasta -d my_custom_db
diamond blastp -q query.fasta -d my_custom_db -o results.tsv
```