Cufflinks error - Processed 0 loci

Question

[wstfljs]

Hi all,

I am trying to assemble a transcriptome using a genome-based approach with tophat2/cufflinks. While tophat2 gives me quite good results (read mapping rate more than 70%) I cannot assemble any transcripts with cufflinks. It doesn't return any specific error, however the number of processed loci equals 0 and the transcripts.gtf file is empty. Below I paste the output of cufflinks and tophat2.

---

tophat_out/align_summary.txt
Left reads:
          Input     :  36870447
           Mapped   :  26922808 (73.0% of input)
            of these:    288345 ( 1.1%) have multiple alignments (0 have >20)
Right reads:
          Input     :  36870447
           Mapped   :  26995453 (73.2% of input)
            of these:    283619 ( 1.1%) have multiple alignments (0 have >20)
73.1% overall read mapping rate.

Aligned pairs:  25007182
     of these:    267231 ( 1.1%) have multiple alignments
                   43433 ( 0.2%) are discordant alignments
67.7% concordant pair alignment rate.

---

You are using Cufflinks v2.2.1, which is the most recent release.
[18:36:04] Inspecting reads and determining fragment length distribution.
> Processed 1 loci.                            [*************************] 100%
> Map Properties:
>    Normalized Map Mass: 3198020.00
>    Raw Map Mass: 3198020.00
>    Fragment Length Distribution: Empirical (learned)
>                  Estimated Mean: 182.67
>               Estimated Std Dev: 59.56
[18:37:19] Assembling transcripts and estimating abundances.
> Processed 0 loci. 

[Devon Ryan]

That seems odd. Can you post the cufflinks commands that you issued? Assuming that you used a GTF or other annotation file with cufflinks, can you confirm that the chromosome names match what's in your BAM files?

[wstfljs]

The command I use is very simple:

cufflinks -p 8 -o cufflinks_out tophat_out/accepted_hits.bam

I expect cufflinks to calculate the fragment length mean and standard deviation on its own, so I don't provide these parameters.

[Devon Ryan]

That should work. If you look at the alignments in a genome browser (e.g., IGV), do you see alignments aggregating into obvious transcripts?

[xiaofeiwang198266]

Hi, did you figure out the reason for the error?

[aleka]

Hi, Did you figure out the error, Seems that I have the same problem.