transcripts are lost after stringtie merge

[Kennyluo4]

Hello, I'm using Stringtie v2.0.4 for my RNAseq data analysis. I have a reference genome and annotation file but I still intend to identify novel genes since the annotation is not complete for my species. I used:

for f in *.bam; do echo ${f}; stringtie -p 8 -f 0.3 -j 5 -G gene_models_main.gff3 -o ${f%.*}.gtf ${f}; done to assemble for individual samples. Then used: stringtie --merge -p 8 -G gene_models_main.gff3 -o stringtie_merged.gtf ./mergelist.txt to get the nonredundant transcript gtf file.

We have characterized novel genes (2 paralogs ) in our species (tetraploid), so I want to ckeck if the two genes are in my data. But I can only find one in the merged gtf file. When I checked the individual assembly, I can find the missed one in some samples and there are reads covered in this region. But in the merged assembly, it's gone. (see figure)

The merged assembly is on the top, and the rest are the individual assemblies. My target trasncript (start with STRG) is assembled in many samples but not in the merged file. Even more strange is that the close reference gene (arahyL5ZR7F) has no coverage and is not assembled in most samples, but it is there. I tried merging without any filtering criteria and without annotation file. But I cannot see that transcript in the merged file. Regards, Ziliang

[LehmannN]

Hello, did you find a solution ?

[kainth-amoldeep]

I have the same issue. Has it been resolved?

[Kennyluo4]

I have the same issue. Has it been resolved?

I wasn't able to solve this issue. Seems Stringtie sticks to the reference strictly. I used a better annotation for the assembly, and there was no issue because the gene model is correctly annotated. Just make sure you have the best annotation for your analysis, or do the de novo assembly.

[kainth-amoldeep]

Thanks @Kennyluo4