Question: Generate SQ Lite database for cummeRbund not working with CuffDiff, and other general questions
0
gravatar for marcuslevitt
23 months ago by
marcuslevitt0 wrote:

Hi everyone,

I am currently a final year undergraduate studying at UoM set with the final yet project of RNA Seq differential expression. I have been using a breast cancer cell line with added E2 and RNA Seq data collected over a time course; 0, 30, 60 & 90 mins. 

I have used the standard workflow: Groomed, TopHat, Cufflinks, Cuffmerge then CuffDiff. When I run for example 0 vs 30 to search for gene differential expression without clicking yes to the generate SQLite the cuffdiff outputs run to completion. However, when I click yes to generate SQLite database for cummeRbund the runs fail to complete.

Anyone have any idea where I am going wrong?

Also if you could help me with these questions that I was be grateful.....

When using cuffmerge to merge cufflinks assembled transcripts output do I need to use a annotated reference, if one was used for the cufflinks input.

The paper used Ilumina HiSeq 2000 but I do not know the insert size and the read length. Does anyone know what the mean inner distance between mate pairs value would be for the Tophat mapping stage?

Many Thanks

 

cummerbund cuffdiff sq lite • 845 views
ADD COMMENTlink modified 23 months ago by Jennifer Hillman Jackson23k • written 23 months ago by marcuslevitt0
0
gravatar for Jennifer Hillman Jackson
23 months ago by
United States
Jennifer Hillman Jackson23k wrote:

Hello,

cummeRbund the runs fail to complete

Is the job running out of resource (memory or walltime)? If memory, then moving to a local or cloud Galaxy with more resource is the solution. If walltime, then you could try running the job on the longer running cluster Stampede (on the tool form under "Job Resource Parameters").

Section 2.8 of the Galaxy support wiki explains alternatives for working with data/jobs that exceed the compute resources at http://usegalaxy.org:
https://wiki.galaxyproject.org/Support

When using cuffmerge to merge cufflinks assembled transcripts output do I need to use a annotated reference, if one was used for the cufflinks input.

This is not required, but the results will be annotated better if you include it (and the annotation has the attributes Cuffdiff utilizes: gene_name, tss_id, gene_id). See the manual for how these attributes are used by Cuffdiff and the other tools: http://cole-trapnell-lab.github.io/cufflinks/manual/

Does anyone know what the mean inner distance between mate pairs value would be for the Tophat mapping stage?

Try the tool NGS: Picard CollectInsertSizeMetrics. Warning: several Picards tools related to this one have recently been problematic at http://usegalaxy.org, so if the tool errors, follow progress on the correction here (or use the tool on a local or cloud Galaxy): https://github.com/jennaj/support-known-issues/wiki

Thanks, Jen, Galaxy team

ADD COMMENTlink written 23 months ago by Jennifer Hillman Jackson23k
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