Finding a gene of interest in contigs from a transcriptome sequencing.

Question

[alexille5640]

Excuse my lack of understanding.

So I have the sequence of a gene of interest I want to locate and compare within the transcriptome of a certain organism (P. leucopus).

The only resource I could find online is here: https://datadryad.org/resource/doi:10.5061/dryad.6hc0f/3 From my understanding the FASTA file from this resource contains a number of contigs.

As I mentioned before I have the sequence of my gene of interest. Now I want to find where it is located within this group data of contigs, so that I can compare my sequence with the one that should be within the transcriptome sequence.

My logic is that this would be similar to CTRL+F function when searching for a phrase within a large document of text.

Where do I begin? What software should i use? Do I open up each contig manually and search for my sequence?

Any help would be greatly appreciated! Excuse my lack of understanding, I am new to the field of bioinformatics.

[Mehmet]

Do you want to know coordinates of your gene in transcriptome of other organism or you want to know homology with the transcriptome? There are several tools (blast, LAST etc) for alignment.

http://last.cbrc.jp

[alexille5640]

Something like that, I would like to locate the best matching sequence in the transcriptome to my sequence.

I have the sequence of a gene of interest. I have a FASTA file containing a set of contigs from a transcriptome sequencing. How can I create a custom database out of this FASTA file so I can BLAST my gene of interest sequence against it?

Assume I'm a complete noob, I'm sorry!

[natasha.sernova]

Try the following orthologous database.

https://omabrowser.org/oma/home/

Select there 'general search'.

Who knows, it may help you.

Answer 1

[lieven.sterck]

Blast is gonna be your tool of choice!

Is your gene sequence (nucleotides?) of the same species as the one of the transcriptome? When yes: use blastn ; if no: use tblastx .

[alexille5640]

It is nucleotides. Alright, so here is what I'm thinking.

I have the sequence of a gene of interest. I have a FASTA file containing a set of contigs from a transcriptome sequencing. How can I create a custom database out of this FASTA file so I can BLAST my gene of interest sequence against it?

Assume I'm a complete noob, I'm sorry!

[Mehmet]

To search gene of interest against the FASTA file using blast locally;

First, create a local database using makeblastdb command of blast as below;

makeblastdb -dbtype nucl -in transcriptome.fa

Then run blastn as below:

blastn -db transcriptome -query yourgenesequences.fa -evalue 1e-5 -outfmt 7

note: evalue that I wrote depends on what you want to know about the alignments. You can modify evalue, outfmt options and/or you can add other parameters.

[GenoMax]

You may want to consider using blat instead (https://genome.ucsc.edu/FAQ/FAQblat ) . It will not require creation of any databases etc. Running a search would be simple. Take a look at the page above and ask questions as needed. You can even ask blat to output search results so they look like a typical blast+ result.

[lieven.sterck]

the same species or other species of my answer is equally (if not more) important ! Do think about what data you have.