This tool assembles RNA-seq reads into transcripts using version Cufflinks 2.2.1.
Given aligned RNA-Seq reads, Cufflinks assembles the alignments into a parsimonious set of transcripts. It then estimates the relative abundances of these transcripts based on how many reads support each one, taking into account biases in library preparation protocols. It is recommended to use the TopHat aligner to map your reads to the reference genome. You can merge the resulting GTFs from several samples to one using the Cuffmerge tool, and use it in differential expression analysis using Cuffdiff.
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.
Cufflinks can perform reference annotation based transcript assembly (RABT) where reference transcripts will be tiled with faux-reads to provide additional information in assembly. If you enable RABT, the output will include all reference transcripts as well as any novel genes and isoforms that are assembled. Please note that RABT can produce false positive novel transcripts when used with complete annotations. Therefore it is recommended only for organisms where deep annotations do not already exist.
The analysis output consists of the following:
This tool uses the Cufflinks package for statistical analysis. Please cite the following article:
Trapnell C, Williams BA, Pertea G, Mortazavi A, Kwan G, van Baren MJ, Salzberg SL, Wold BJ, Pachter L. Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat Biotechnol. 2010 May;28(5):511-5.
If you use RABT, please cite
Adam Roberts, Harold Pimentel, Cole Trapnell, Lior Pachter. Identification of novel transcripts in annotated genomes using RNA-seq. Bioinformatics 2011. 27 (17): 2325-2329.