creating a miRNA gtf by mirdeep2

Question

[zizigolu]

hi,

how I can create my own annotation gtf for miRNA DE analysis by mirdeep2?

thank you

[WouterDeCoster]

I'm not sure about mirdeep2, but there is something else you could try. If I'm not mistaken, based on your other posts, miRNA annotation is not available for your species of interest. A possibility might be to map all the reads (you probably already did that?), convert bam files to bed files (e.g. bedtools), merge all bed files (bedtools) and create your own annotation file... It could be used for differential expression analysis, but interpretation would be harder. But it would allow you to identify which coding elements are differential expressed between your two conditions. Investigating the function of differential expressed small RNAs (perhaps in other species which richer annotation) is then a next step...

[zizigolu]

thank you for paying attention. yes in mirbase there is not miRNA annotation. in ensembl gff3 contains miRNA but when I converted that to gtf, tophat gave me error

[WouterDeCoster]

I'm not sure if you posted it already elsewhere, but if you can be more specific about that error then that's perhaps something that can be fixed. Also, a gtf isn't absolutely required for mapping...

[zizigolu]

thank you. I will post errors. but I need gtf for featurecounts or cufflinks

[WouterDeCoster]

Definitely. But if your annotation is poor, you can create your own gtf as described above based on all reads in all samples combined.

[zizigolu]

sorry I converted my gff3 to gtf but there is no more miRNA in my resulted gtf

head(gtf[,1:9])

I JCVI exon X216 X836 . X. ..1

1 I JCVI CDS 216 836 . + 0

2 I JCVI exon 3952 4253 . + .

3 I JCVI exon 4774 4963 . + .

4 I JCVI CDS 3952 4253 . + 0

5 I JCVI CDS 4774 4963 . + 1

6 I JCVI exon 6805 6881 . - .

[zizigolu]

sorry if I use gff3 contains miRNA from ensembl for featurecounts, is this something like I have used miRNA spesific annotation like which there are in mirbase or this gff3 is different and DE is not proper?

Answer 1

[John]

Congratulation Angel for getting the rarest badge on all of Biostars! :)

[zizigolu]

thank you John :) :) :)