Cuffdiff Error - sort order of reads in BAMs must be the same

Question: Cuffdiff Error - sort order of reads in BAMs must be the same

3.9 years ago by

United States

Hi,

I am trying to run Cuffdiff with a Cuffmerged data of 8 files along with their Tophat counterparts and am coming up with an Error "sort order of reads in BAMs must be the same". Will be awesome if I can get some guidance to get this issue fixed.

Cheers

Francis

rna-seq cuffdiff • 3.6k views

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modified 21 months ago by takshan • 10 • written 3.9 years ago by francis.gandhi • 20

3.9 years ago by

Jennifer Hillman Jackson ♦ 25k

United States

Jennifer Hillman Jackson ♦ 25k wrote:

Hello,

It could be the ordering of the data in the input reference dataset (GTF or GFF3). This workflow can be used to sort it correctly for this tool. All steps that use it will need to be executed again (Cufflinks, if used there - and Cuffmerge).
http://usegalaxy.org/u/jeremy/p/transcriptome-analysis-faq#faq5

The source does not need to be Ensembl, but the reference genome the GTF/GFF3 data is based on must be an exact match for the reference genome specified with any tool (if/when used). Meaning, the chromosome identifiers must be exactly the same. In Cuffdiff, this is used when "Bias correction" is checked. Avoid that option if the reference genome is not a match for one in Galaxy and you are not providing it as a Custom Reference Genome. How to check versus Galaxy native genomes or add a Custom reference:
http://wiki.galaxyproject.org/Support#Reference_genomes
http://wiki.galaxyproject.org/Support#Custom_reference_genome

Thanks, Jen, Galaxy team

ADD COMMENT • link written 3.9 years ago by Jennifer Hillman Jackson ♦ 25k

Thank you so much for the information Jennifer. I will give it a shot.

Cheers, Francis

ADD REPLY • link written 3.9 years ago by francis.gandhi • 20

Thanks Francis! If this works, it would be great if you could "accept" the answer so that others may easily find it. Just click on the checkmark next to the answer. Much appreciated! Jen

ADD REPLY • link written 3.9 years ago by Jennifer Hillman Jackson ♦ 25k

21 months ago by

takshan • 10

India

takshan • 10 wrote:

Am also having the same issue. am unable to RUN the workflow provided in FAQ to sort also using sort sam/bam am unable to use in cuffdiff. Any guidance would be of great help.

ADD COMMENT • link written 21 months ago by takshan • 10

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