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Kallisto Quant Specifications

Process Details

  • Name: kallisto_quant
  • Process UUID: zjmzdeecFmXy1rNlBUO6F7EaQNzwCM
  • Process Group: Transcript Quant.

Overview

Kallisto quant runs the quantification algorithm that computes equivalence classes for reads and quantifies transcript abundances. This process performs pseudo-alignment-based RNA-seq quantification, providing fast and accurate transcript-level abundance estimates without the need for full genome alignment.

This process is implemented in Bash, which invokes Python scripts for GTF filtering and various bioinformatics tools including kallisto, gffread, and samtools for comprehensive transcript quantification and BAM file processing.

Key Functionality

  • Transcript Quantification: Performs pseudo-alignment and abundance estimation using kallisto's probabilistic model
  • GTF Processing: Filters and reformats GTF annotation files to ensure compatibility with genome sequences
  • BAM Generation: Optionally projects pseudoalignments to genome-sorted BAM files for visualization and downstream analysis
  • Quality Control: Processes and sorts BAM files while removing unmapped reads for clean output

Input/Output Specification

Inputs

Required Inputs

  • Mate Information

    • Description: Specifies whether reads are single-end or paired-end
    • Format: Value specification

  • Reads

    • Description: RNA-seq reads in FASTQ format for quantification
    • Format: FASTQ files

  • Kallisto Index

    • Description: Pre-built kallisto transcriptome index for pseudo-alignment
    • Format: IDX file

  • GTF File

    • Description: Gene annotation file containing transcript and exon coordinates
    • Format: GTF file

  • Genome Sizes

    • Description: Chromosome size information for BAM file generation
    • Format: Sizes file

  • Genome

    • Description: Reference genome sequence in FASTA format
    • Format: FASTA file

Outputs

  • Output Directory

    • Description: Directory containing kallisto quantification results including abundance estimates
    • Format: Directory

  • BAM Files

    • Description: Genome-aligned BAM files generated from pseudoalignments (when genomebam option is enabled)
    • Format: BAM files

  • BAM Index

    • Description: Index files for the generated BAM files to enable rapid access
    • Format: BAM index files

Parameters & Settings

These parameters can be adjusted in the Foundry UI when running this process.

  • Single or Paired End Reads

    • Description: When single is selected, the fragment_length and standard_deviation parameters are REQUIRED.
    • Available options: single, pair (default)

  • Fragment Length

    • Description: Estimated average fragment length. Required when using single reads. Typical value: 200
    • Default value: 200

  • Standard Deviation

    • Description: Estimated standard deviation of fragment length. Required when using single reads. Typical value: 30
    • Default value: 30

  • Kallisto Parameters

    • Description: Kallisto parameters.
    • Default value: --threads 4

  • Genome BAM

    • Description: If true is selected, Kallisto will project pseudoalignments to genome sorted BAM file.
    • Available options: true (default), false

References & Resources

  • Tool Documentation: Contact the team for details on filter_gtf_for_genes_in_genome.py
  • Related Papers: Bray, N.L., Pimentel, H., Melsted, P. & Pachter, L. Near-optimal probabilistic RNA-seq quantification. Nature Biotechnology 34, 525–527 (2016). https://doi.org/10.1038/nbt.3519