The applications listed here are available for use in the Discovery Environment and are documented in: Discovery Environment Manual.

Discovery Environment Applications List

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Rationale:

SplAdder, short for Splicing Adder, a toolbox for alternative splicing analysis based on RNA-Seq alignment data. Briefly, the software takes a given annotation and RNA-Seq read alignments in standardized formats, transforms the annotation into a splicing graph representation, augments the splicing graph with additional information extracted from the read data, extracts alternative splicing events from the graph and quantifies the events based on the alignment data, the quantified events can then be used for differential analysis. SplAdder in its default configuration, consists of the following steps:

  • transform annotation into splicing graph representation
  • generate an augmented splicing graph for each alignment file by inferring and adding the following elements:
    • insert intron retentions
    • insert cassette exons
    • insert new intron edges
  • merge the augmented splicing graphs into a common splicing graph
  • quantify all alternative splicing events on each of the provided alignment files

SplAdder is invoked using the following:

  1. Input (s)
    1. The folder containing the list of alignment files (bam files)
    2. The genome annotation file (gtf file)
  2. Parameters
    1. Alternative Splicing event type (Default is Exon_skip, Intron_retention, alternate_3',alternate_5', multiexon_skip):
      • Exon_skip
      • Intron_retention
      • alternate_3'
      • alternate_5'
      • multiexon_skip
    2. Confidence level (Default is 3)
    3. Primary_alignments_only (Default is no)
    4. Merge Strategy (Default is merge_graphs)
  3. Outpus
    1. An output directory where results files are stored (Default is Output). 

Please work through the documentation and add your comments on the bottom of this page, or email comments to support@cyverse.org. Thank you.

Test Data

 The test data for Spladder-1.0.0 consists of 6 bam files and an annotation files from Arabidopsis thaliana 

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All files are located in the Community Data directory of the iPlant Discovery Environment at the following path: Community Data > iplantcollaborative > example_data > spladder (/iplant/home/shared/iplantcollaborative/example_data/spladder/)

Run SplAdder as following:

  1. Input (s)
    1. The folder containing the list of alignment files - /iplant/home/shared/iplantcollaborative/example_data/spladder/
    2. The genome annotation file (/iplant/home/shared/iplantcollaborative/example_data/spladder/TAIR10_GFF3_genes.tiny.gff)
  2. Parameters
    1. Alternative Splicing event type - Exon_skip, Intron_retention, alternate_3',alternate_5', multiexon_skip
    2. Confidence level - 3
    3. Primary_alignments_only - no
    4. Merge Strategy - merge_graphs
  3. Outputs
    1. Output folder containing the following files
      merge_graphs_alt_3prime_C3.confirmed.pickle
      merge_graphs_alt_3prime_C3.counts.hdf5
      merge_graphs_alt_3prime_C3.pickle
      merge_graphs_alt_5prime_C3.confirmed.pickle
      merge_graphs_alt_5prime_C3.counts.hdf5
      merge_graphs_alt_5prime_C3.pickle
      merge_graphs_exon_skip_C3.confirmed.gff3
      merge_graphs_exon_skip_C3.confirmed.pickle
      merge_graphs_exon_skip_C3.confirmed.txt
      merge_graphs_exon_skip_C3.counts.hdf5
      merge_graphs_exon_skip_C3.pickle
      merge_graphs_intron_retention_C3.confirmed.pickle
      merge_graphs_intron_retention_C3.counts.hdf5
      merge_graphs_intron_retention_C3.pickle
      merge_graphs_mult_exon_skip_C3.confirmed.pickle
      merge_graphs_mult_exon_skip_C3.counts.hdf5
      merge_graphs_mult_exon_skip_C3.pickle

merge_graphs_exon_skip_C3.confirmed.txt contains the following output

 

Tool Source for App