Differential expression using Cuffdiff


This tool performs differential expression analysis using the Cufflinks package version 2.1.1.

You will need to provide a list of file names of replicate BAM files belonging for each group. These lists can be generated with the tool Utilities / Make a list of file names. The file names of these lists are used to set the labels in the output files, so they can be named something relevant to your use case, e.g. treatment.txt and control.txt.



Given GTF and BAM files, Cuffdiff performs differential expression analysis of genes and transcripts using the Cufflinks algorithm.

Cufflinks can detect sequence-specific bias and correct for it in abundance estimation.

By default, Cufflinks will uniformly divide each multi-mapped read to all of the positions it maps to. If multi-mapped read correction is enabled, Cufflinks will re-estimate the transcript abundances dividing each multi-mapped read probabilistically based on the initial abundance estimation, the inferred fragment length and fragment bias, if bias correction is enabled.

FDR-adjusted p-values (q-values) are calculated. The concise output files include only those genes or transcripts which have a q-value lower than the given FDR. The value of the Significant-column is adjusted accordingly (yes/no) in all output files.


The concise analysis output consists of the following files:

If the complete output is selected, the following optional outputs may also be generated:

1. FPKM tracking files

2. Count tracking files

3. Read group tracking files

4. Differential expression tests

5. Differential splicing tests

6. Differential coding output

7. Differential promoter use

8. Read group info

8. Run info


This tool uses the Cufflinks package for statistical analysis. Please read the following article for more detailed information:

Trapnell et al. Differential analysis of gene regulation at transcript resolution with RNA-seq. Nat Biotechnol. 2013 Jan;31(1):46-53.