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Sequential Mapping Specifications

Process Details

  • Name: Sequential Mapping
  • Process UUID: 20bdb83c-ed05-11e8-9ce4-85221b21564c
  • Process Group: Alignment

Overview

Sequential Mapping is a comprehensive alignment process designed to filter out and quantify specific sequence sets from sequencing data. The process performs sequential mapping against multiple reference databases, allowing for the systematic removal of specific RNA types (such as rRNA, miRNA, tRNA, snRNA, snoRNA) or repetitive elements (rmsk) from the dataset. This approach enables researchers to progressively filter their data and focus on sequences of interest while maintaining detailed mapping statistics for each step.

This process is implemented in Bash, which invokes multiple alignment tools including Bowtie, Bowtie2, and STAR for sequence mapping and various bioinformatics utilities for post-processing.

Key Functionality

  • Multi-aligner Support: Supports Bowtie, Bowtie2, and STAR aligners with customizable parameters for each mapping step
  • Sequential Filtering: Progressively maps and filters reads against multiple reference databases in a defined order
  • Duplicate Removal: Optional removal of PCR and optical duplicates using either Picard MarkDuplicates or UMI-based deduplication
  • Strand-specific Analysis: Generates separate sense and antisense alignment files when requested
  • Visualization Support: Creates bigWig files for genome browser visualization
  • Comprehensive Statistics: Provides detailed mapping statistics and quality metrics for each alignment step

Input/Output Specification

Inputs

Required Inputs

  • Sequencing Reads

    • Description: FASTQ files containing sequencing reads to be mapped
    • Format: FASTQ (compressed or uncompressed)

  • Mate Type

    • Description: Specifies whether the data is single-end or paired-end
    • Format: Mate specification value

Optional Inputs

  • Common Database

    • Description: Directory containing common RNA sequence databases (ercc, rRNA, miRNA, tRNA, snRNA, snoRNA, rmsk)
    • Format: Directory

  • Bowtie Index

    • Description: Bowtie alignment index files
    • Format: Bowtie index files

  • Bowtie2 Index

    • Description: Bowtie2 alignment index files
    • Format: Bowtie2 index files

  • STAR Index

    • Description: STAR alignment index files
    • Format: STAR index files

Outputs

  • Filtered Reads

    • Description: Remaining unmapped reads after sequential filtering
    • Format: FASTQ

  • Alignment Statistics

    • Description: Detailed mapping statistics for each alignment step
    • Format: BOW log files

  • BAM Alignment Files

    • Description: Sorted and indexed BAM files for each mapping step
    • Format: BAM with corresponding BAI index files

  • Log Files

    • Description: Process execution logs and alignment statistics
    • Format: LOG

  • BigWig Files

    • Description: Genome browser visualization files (optional)
    • Format: BigWig

Parameters & Settings

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

  • Remove Duplicates

    • Description: Duplicates (both PCR and optical) will be removed from alignment file (bam) and separate count table will be created for comparison
    • Available options: no (default), yes

  • Remove Duplicates Based on UMI After Mapping

    • Description: UMI extract process should have executed before this step. Read headers should have UMI tags which are separated with underscore.(eg. NS5HGY:2:11_GTATAACCTT)
    • Available options: no (default), yes

  • Create BigWig

    • Description: Create bigWig file for IGV
    • Available options: no (default), yes

  • Remove Previous Reads

    • Description: After successful completion of the sequential mapping, previous reads will be removed to save space. However, this might affect resume functionality.
    • Default value: false

References & Resources

  • Tool Documentation: Contact the team for details on alignment and post-processing scripts
  • Related Papers:
  • Langmead B, Salzberg SL. Fast gapped-read alignment with Bowtie 2. Nature Methods. 2012;9(4):357-359. doi:10.1038/nmeth.1923
  • Dobin A, Davis CA, Schlesinger F, et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics. 2013;29(1):15-21. doi:10.1093/bioinformatics/bts635