I have a project where I need to theorize the protein sequence of quail myoglobin. A very similar organism (chicken) has already had its myoglobin sequenced. I can access the entire genome of quail online. I can also access the gene nucleotide sequence and amino acid sequence of chicken myoglobin online as well.
I've been trying to use the tools on BLAST online to try and find similar DNA sequences to the chicken myoglobin in quail but I am having significant difficulties
I think I would recommend to make your own BLAST database for quail.
You can download the BLAST software from NCBI (search for 'local blast' in your search engine of choice).
Once you have the software, one of the commands is to make your own BLAST database - you can use either the genome sequence of quail (in a FASTA file) or a file of all the CDS from quail, to make your database.
Once you have the database made you can use the local BLAST command to search for the chicken protein sequence in the quail genome, using the tBLASTn function.
Once you have found the best matching quail genomic region corresponding to the chicken protein, you can work out how to translate that nucleotide sequence into protein sequence.
Hi,
I quickly had a look on quail databases... it looks poor, what database do you use ?
As Jonathan wrote, you can make your own BLAST database.
But, according to what I looked about quail databases, this will be propbaly a bit of work because genomic sequence of quail is poorly assembled (japanese quail).
I just tested some basic analysis using online BLAST on NCBI website. I think you can manage to get nucleotidic and protein sequences of quail myoglobin :
1) by aligning (using BLASTn) nucleotid sequence of chicken to quail's genome sequence (wgs database on NCBI) then according to the test I did, you will have 4 contigs of interest (Coturnix japonica DNA, contig: contig_137770, Coturnix japonica DNA, contig: contig_137773, Coturnix japonica DNA, contig: contig_137775, Coturnix japonica DNA, contig: contig_137776).
2) by aligning (using tBLASTn) protein sequence of chicken to quail's genome sequence (wgs database on NCBI). You will have 3 contigs of interest (Coturnix japonica DNA, contig: contig_137770, Coturnix japonica DNA, contig: contig_137773, Coturnix japonica DNA, contig: contig_137775).
By assembling these sequences, I think you will be able to have the nucleotid sequence of quail myoglobin. And by translating the assembled sequence, you will have the protein sequence.
Good luck,
me again,
I tried to retrieve the myoglobin sequence of quail (Coturnix japonica) using the sequence of gallus gallus myoglogin (genBank accession : NM_001167752)... The results I had are in the attached file.
It worked not so bad.Just a short part of the sequence couldn't be found using GenBank databases, but I guess this can be done by a deeper analysis.
I hope this will help you
Bye
Thank you for this. You have corroborated the results I produced by creating a local database from the wgs data. I was becoming frustrated because I had been using an early draft database locally, and online I had been searching with those bad assemblies you had mentioned (especially because earlier this year I had downloaded and been using the wgs database without realizing it and getting the appropriate results).
Part of that short missing sequence, NDMASKY, is actually one of the few pieces I have been unable to verify via mass spec. I am currently running glu-c digests to reveal it. Would you happen to know if i ran local searches on some of the raw data (apparently available here: http://www.ncbi.nlm.nih.gov/sra/?term=DRA000595) if I might be able to predict the missing sequence? There are several very large files I could convert to fasta that I wonder might have what I need (I am very unfamiliar with these database systems and bioinformatics as a whole).
-Many thanks
Hi John. I manage to partially analyse the raw data DRA000595 with online tool (link below) ... I'm not sure it is necessary to build a so large local database, just for this.
http://blast.ncbi.nlm.nih.gov/Blast.cgi?PROGRAM=blastn&PAGE_TYPE=BlastSearch&PROG_DEFAULTS=on&BLAST_SPEC=SRA
Here is a list (non exhaustive) of reads sequences that seems to confirm that the short missing sequence is 100% similar to the chicken one (see attached file).
Hope this will help
Good luck, bye
Ah, yes that was much easier than making a local db (which I tried but ran into a few issues). Thanks for the assistance.
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