RNA-SEQ reads using flux-simulator

Question

[madiha_shabbir46]

Hi,

I am having some issues with flux-simulator, I am using it for the first time to generate reads from my reference transcriptome sequence. It requires gtf of my reference, so I made a one using my own script (couldn't find any other tool to do this for me) according to the format mentioned at this site http://confluence.sammeth.net/display/SIM/.GTF+Gene+Annotations

but I get the error to add transcript_id too which is not mentioned where to add, so I tried each column but still get different kinds of errors.

 Error in line 41667: invalid score 'exon'.
 Error in line 41668: invalid score 'exon'.
 Error in line 41669: invalid score 'exon'.
 Error in line 41670: invalid score 'exon'.
 Error in line 41671: invalid score 'exon'.
     OK (00:00:00)
found 0 transcripts

[PROFILING] Parameters
NB_MOLECULES    0
EXPRESSION_K    -0.6
EXPRESSION_X0   9500.0
EXPRESSION_X1   9.025E7
PRO_FILE_NAME   /media/madiha/Data1/DTR/softwares/flux_simulator/PAR.pro
profiling  OK (00:00:00)
Updating .pro file   OK (00:00:00)
molecules   0
[ ERROR] Profiler has no molecules!
java.lang.RuntimeException: Profiler has no molecules!
at barna.flux.simulator.SimulationPipeline.call(SimulationPipeline.java:438)
at barna.flux.simulator.SimulationPipeline.call(SimulationPipeline.java:54)
at barna.commons.launcher.Flux.main(Flux.java:198)

It would be so kind if someone can share his/her sample files for flux-simulator.

Regards,

Madiha

Answer 1

[Istvan Albert]

Looks like the GTF file that you made is incorrect.

The score should be in column 6 and just set that to 0. What you have there (exon) should be in column 3 called type.

Make a short GFT file with just ten lines or so, look at it in an editor to ensure it is correct. Simulate with that. If that works then it is all about ensuring your GFT stays correct for all other lines.