![]() ![]() The module can optionally perform this mapping for you. While you can open your GTF to look up these references, you may find it more convenient to have them mapped to gene symbols in the GCT so that these are available in any downstream processing. Each line of the GTF will contain several named attributes that show the tracking_ids associated with that particular feature. These tracking_ids refer to entries in the GTF file supplied to Cuffdiff and will take the form of XLOC_00001 for genes, TCONS_00000001 for isoforms, TSS1 for tss references, and P1 for cds references. GTF Name MappingÄepending on your prior workflow, your read_group_tracking files may contain internal Cufflinks tracking_id references rather than gene symbols this is particularly the case if you used the merged.gtf file from Cuffmerge as an input to Cuffdiff. See this section of the Cufflinks manual for a full explanation of the read_group_tracking format, which is the source of the above description. Note that doing so requires that all values for the entire feature be screened - even those with OK status - rather than just those for affected replicates. The default is to screen as most downstream modules cannot handle "NA" values. In practical terms, any value associated with a non-OK status represents a quantification error and so will be either screened out of the resulting GCT or else replaced with an "NA". FAIL: when an ill-conditioned covariance matrix or other numerical exception prevents deconvolution.LOWDATA: too complex or shallowly sequenced.Within the read_group_tracking file, Cuffdiff gives a status for the quantification calculation result per tracking ID and replicate: ![]() FPKM (the default): FPKM of this object in this replicate.external_scaled_frags: Estimated number of fragments originating from the object, after transforming to the external common count scale (for comparison between conditions).internal_scaled_frags: Estimated number of fragments originating from the object, after transforming to the internal common count scale (for comparison between replicates of this condition.).raw_frags: The estimate number of (unscaled) fragments originating from the object in this replicate.The module will extract FPKM values by default, but it can also extract any of the other available types of quantification values as well: Cuffdiff stores these in several read_group_tracking files at the level of genes, isoforms, transcription start sites (tss_group), and coding sequences (cds), each named appropriately for the feature information it holds. This provides independent FPKM and other quantification values for each replicate of each condition in the dataset, which are thus suitable as input to existing GenePattern modules for analysis and visualization such as ConsensusClustering, NMFConsensus, and so on. The Cuffdiff module performs a quantification step as a precursor to running its differential expression calculations. This module will convert files in the Cufflinks read_group_tracking format into GenePattern's GCT format for use in downstream modules. ![]()
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