INSA - TP bioinformatique

Exercise 5: Database against database BLAST searches

Objective: compare the proteomes of Mycoplasma genitalium and Mycoplasma pneumoniae.

Step 1: Install BLAST on your local computer. BLAST executables are available at ftp://ftp.ncbi.nih.gov/blast/ (or HERE ). Carefully read the instructions in the BLAST distribution to install all necessary files and executables in your environment.

Create a directory named blast, and then de-compress and de-tar the BLAST archive in this directory.

Move BLAST executables (blastall and formatdb) in your bin directory.

Create in your home directory a file named .ncbirc. This file must contain the following lines (replace /home/duret/blast/ by the path to your own blast directory):

[NCBI] 
Data="/home/duret/blast/data/"

Define the following environment variables:

set BLASTDB=/home/duret/blast/db/ (default repertory for BLAST databases)

set BLASTMAT=/home/duret/blast/matrix/ (repertory containing substitution matrices)

More help.

Step 2: For each species (Mycoplasma genitalium, Mycoplasma pneumoniae), select all protein entries from SwissProt-TrEMBL using WWW-Query at PBIL, and then extract sequences in FASTA format.

How many proteins are encoded by Mycoplasma genitalium and by Mycoplasma pneumoniae genomes ?

Step 2.1

Databank: SwissProt-TrEMBL

Species: Mycoplasma genitalium

Submit, and then retrieve sequences in FASTA format.

mg.seq

Step 2.2

Databank: SwissProt-TrEMBL

Species: Mycoplasma pneumoniae

Submit, and then retrieve sequences in FASTA format.

mp.seq

Step 3: Prepare a BLAST database with Mycoplasma pneumoniae proteins, and then BLAST (blastp) Mycoplasma genitalium proteins against this database.

formatdb -i mp.seq
blastall -p blastp -d mp.seq -i mg.seq -o mg.blast  -e 0.001

mg.blast

How many Mycoplasma genitalium proteins have at least one homologue in Mycoplasma pneumoniae ?

blastall -p blastp -d mp.seq -i mg.seq -o mg.blast2 -e 0.001 -v 1 -b 1 -m 8
wc mg.blast2

mg.blast2