getSpliced() error: improper genomic coordinate 46954453 on chrUn_random for TCONS_00084296

I have searched through all the threads on this topic and I have yet to find a reasonable explanation for why I continue to encounter this error. I have already run Cufflinks on my samples, but when I attempt to run Cuffmerge (cuffdiff -o /scratch/sbag/Tikshana/BANANA/cuff/Macbal_diff -b /scratch/sbag/Tikshana/BANANA/RAW/Mac.fa -p 8 -u /scratch/sbag/Tikshana/BANANA/cuff/merged_asm/merged.gtf -L leaf,root /scratch/sbag/Tikshana/BANANA/map/Top_Macbal_6127/accept
ed_hits.bam, /scratch/sbag/Tikshana/BANANA/map/Top_Macbal_6141/accepted_hits.bam), I get the following error:

Error (GFaSeqGet): end coordinate (46954453) cannot be larger than sequence length 46622217
GffObj::getSpliced() error: improper genomic coordinate 46954453 on chrUn_random for TCONS_00084296


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This problem looks like a genome mismatch issue. Data was mapped against one version of the genome but the annotation was based on a different one? My guess is both happened to have a “chrUn_random” chromosome but they are different lengths, which means different content. All of the coordinates in your data will be wrong if this is true – and the tool will not necessarily error, just produce scientifically incorrect results.

That said, all of these tools are deprecated when used in Galaxy, as explained in your other post here:

Why and the alternatives are covered in this prior Q&A:

If you want to try to troubleshoot line-command usage, there was a google group – I don’t think it is very active anymore but you may find prior help. Biostars, Seqanswers, and other general bioinformatics forums will also have much prior discussion. These are usually common enough that even just a basic web search with the error message quoted will find help.

But avoid using the tools in Galaxy, or expected problems.


Solved by checking all paths and file name, one negligible mistake might abandon the whole experiment.

Make it simple by checking all file name and location is deeply suggested