Ref seq for mapping

Question

[Arash]

Hi again ,

How can I set or download Chicken ref and annotated seq as Ref for mapping?

[Farbod]

Hi, have a look at

https://www.ncbi.nlm.nih.gov/genome/?term=chicken%5Borgn%5D

and

ftp://ftp.ensembl.org/pub/release-86/fasta/gallus_gallus/dna/

Answer 1

[GenoMax]

You can find sequence/annotation/aligner index bundles for chicken sequence at Illumina iGenomes site.

If that site does not work from Iran then use the Ensembl chicken genome site to get sequence/annotation. You will need to build your own aligner indexes.

[Arash]

Dear genimax2,

Thanks so much for your wonderful help.In iGenomes website, I find 2 files in NCBI for Gallus Gallus, build2.1 & build3.1 . I have to download both or??? If both , I have to map with both ...sorry for my stupid question

And the files in NCBI and other sources ( Ensembl or USCS) are different ?

[GenoMax]

Genomes come in different builds. Think of them as releases (like software). Higher number means a newer release. So 3.1 would be a newer/latest build. If you don't have a specific reason to use an older one then always go with the latest. In fact Ensembl site has even a newer release. You should choose and stick with one release. All your analyses would be tied to this release and you would make a note of this in your publication (so others can replicate your results).

Sequence (for a particular build) will not be different (at Ensembl/UCSC) but the annotation may be since the individual sites do more or less or use different processes for annotation.

[Arash]

Thanks,so I just download the build3.1 or bulid2.1 and use as a ref?

[GenoMax]

You need to decide that based on your needs. Like I said above you need only one. Choose one build from one site (Ensembl/UCSC/NCBI) and stick with it for all analysis.

iGenomes bundles have the advantage that they contain all you need to get started right away and there will be no issues with identifiers etc. If you are a new user then you may like that but it does not offer any other advantage.

[Arash]

Hi,

I downloaded the gallus gallus data from iGenome (Zip file).It contains some folders genome, chromosomes ,Bowtie index,Botie2index,..... Which folder or data have to be used as a ref?

[GenoMax]

Reference in fasta format should be in Gallus_gallus/NCBI/genome_build/Sequence/WholeGenomeFasta/genome.fa. This is the basis for all indexes that are included in the download so you do not need to rebuild the indexes (unless you are planning to use an aligner that is not included in the index bundle e.g. BBMap).

[Arash]

Dear genomax2, For RNA seq , where I have to be use ref genome with mRNA annotation?

[GenoMax]

You can align to the genome and then count only those reads that overlap "known" genomic features you are interested in (e.g. exons) that are in the GTF file.

[Arash]

I did map reads (Case / Control) to Gallus Gallus genome Fasta file (iGenome) by Geneious software.The software produce31 map files.When I want to calculate expression level , but I got the following message : You have to use ref file with mRNA annotation" Please help me to select correct ref file.Thanks.

[GenoMax]

If you used the iGenomes fasta then you can use the corresponding GTF file that you can find in Gallus_gallus/NCBI/genome_build/Annotation/Genes/gene.gtf

[Arash]

Hi,Thanks...When I did map with Geneious, we got unmatched reads,Whats the problem?

[GenoMax]

You will always have some unmatched reads. What % of reads was unmatached?

[Arash]

Thanks!I am working on it