Problem with blastp when blasting against custom made database

Question

[thjnant]

Hello,

I have been trying the last days to make my database for blast work but I really don't know how to figure out this problem.

Begining from the start of the problem, I want to make a blastp against a protein database that I have created.

Here is an example of my sequence identifiers and how they are formatted:

>TRA_G345655|locus=scaffold45|99209|101155|-|translate_table|standard

I try to make the blast database with the following command:

makeblastdb -in database.fa -title database -dbtype prot -out protdb -parse_seqids

It does not work and it gives me the following error:

No volumes were created because no sequences were found.

BLAST Database creation error: Defline lacks a proper ID around line 1

When I try it without '-parse_seqids', it works fine and it generated the three files with pin, psq and phr.

I put these 3 files with my database.fa fasta file in one folder called DB and then I try to run blastp with the following command:

blastp -query query.fa -db DB -out proteins_blastp_1e-30_table.txt -evalue 1e-30 -outfmt 6

or with:

blastp -query query.fa -db DB/database.fa -out proteins_blastp_1e-30_table.txt -evalue 1e-30 -outfmt 6

or with:

blastp -query query.fa -db database -out proteins_blastp_1e-30_table.txt -evalue 1e-30 -outfmt 6

But it gives me the following error:

BLAST Database error: No alias or index file found for protein database [database] in search path

How can I solve this problem or where is the origin of this problem?

[Michael]

The scenario you are describing can never happen, blastp would never ask for a nucleotide database, so you first need to get the association between commands and error message right. Most likely that you simply mixed up the blast programs (blastp/blastn/x)

[thjnant]

No, I have not mixed up blastp/blastn/x. This is the simply the situation I am describing, I have two fasta files of proteins and I want to use blastp and this is the error I get which I have copied from my linux shell.I ran the command again, I updated the error, it must have been a mistake with nucleotide.

[Michael]

Yes you have, but you have also updated your question with a new error message, because in the first version it was saying :"No alias or index file found for nucleotide database" now it says protein.

[vigneshprbh37]

Hey can you give me some advice on a blast installation matter?

So for the past two days I have been trying to install and execute a stand alone blast named ncbi-blast-2.2.30+ on a centos os system

i managed to download a nr ref sequence from ncbi ftp using the command

wget ftp://ftp.ncbi.nlm.nih.gov/blast/db/FASTA/nr.gz

and extracted the file using the command

tar -xvpf nr.gz

I got an nr file of 33 gb size

but when I try to format the file using the command

./makeblastdb \
  -in /home/Desktop/ncbi-blast-2.2.30+/db/nr \
  -dbtype 'nucl' \
  -input_type 'fasta' \
  -out /home/Desktop/ncbi-blast-2.2.30+/output

I get the error showing

BLAST Database creation error: FASTA-Reader: No residues given

Can you give any suggestions on the nature of the problem and how I can solve it?

Thanks

Answer 1

[Michael]

The documentation on how to build proper sequence ids is here

In short: your fasta defline doesn't contain a valid identifier and needs to be changed to be parsable. A valid identifier consists of two or three fields separated by |.

E.g. for a ref seq entry:

ref|NM_010450.1|

For your case most likely the sequence has a local identifier, then you should change your defline to start with:

lcl|TRA_G345655

The second error is caused by:

makeblastdb -in database.fa -title database -dbtype prot -out protdb -parse_seqids

so you also need to change this:

blastp -query query.fa -db protdb -out proteins_blastp_1e-30_table.txt -evalue 1e-30 -outfmt 6

[thjnant]

Thank you so so so much, I changed the header and also applied your second comment in the blastp command, and now it is working, perfect. And well, then yes, I had mixed up those things :) Thank you so much again.