StringTie error generating gene and transcripts counts files

[tanvirai]

Hello. I’ve been running tools on Galaxy today to analyze bulk RNA-seq data following this pipeline (https://youtu.be/KVh98S89yUU?si=jvBNvbZ-stVoz9-k). I’ve run StringTie on two different datasets and I keep getting this error while trying to generate the Gene and Transcript counts files.

---

Dataset Error Report

An error occurred while running the tool toolshed.g2.bx.psu.edu/repos/iuc/stringtie/stringtie/2.2.2+galaxy0. Details

Tool generated the following standard error:

/usr/local/bin/prepDE.py:72: SyntaxWarning: invalid escape sequence ‘-’
RE_COVERAGE=re.compile(‘cov “([-+\d.]+)”’)
/usr/local/bin/prepDE.py:75: SyntaxWarning: invalid escape sequence ‘-’
RE_GFILE=re.compile(‘-G\s*(\S+)’) #assume filepath without spaces…

---

I have sent a bug report to the Galaxy administrator but while I wait for their reply, I was wondering if anybody else has faced the same problem or if somebody has some answers. Is this an issue with my dataset or with the StringTie tool?

[jennaj]

Hi @tanvirai

This help will apply for you, too.

@jennaj

The instructions

@igor

A proper workflow is:

1. create gene models for every sample
2. merge individual gene models into a single “master” annotation using StringTie merge
3. and use the master annotation for read counting

The reads can be counted with StringTie or any other software, for example, featureCounts. For read counting with StringTie disable gene prediction.

Read counting should be done with the same annotation, for sake of comparison, hence, the two options were made incompatible.

More details can be found in this other Q&A

@gallardoalba

a detailed explanation of the necessary steps can be found in:

training.galaxyproject.org

Galaxy Training: De novo transcriptome reconstruction with RNA-Seq

Training material for all kinds of transcriptomics analysis.

Cheers.

---

Right now, you are calling counts against a single output from a single Stringtie run (scientific problem), and the option to call new gene models is still on (technical problem).

Hope this helps, and thanks for sending in the bug report for supporting data, very helpful!

[tanvirai]

Thank you!!