mapping.md

Mapping

Introduction

The mapping pipeline has been created for NGS users who want to align there data with the most commonly used alignment programs. The pipeline performs a quality control (QC) on the raw fastq files with our Flexiprep pipeline. After the QC, the pipeline simply maps the reads with the chosen aligner. The resulting BAM files will be sorted on coordinates and indexed, for downstream analysis.

Tools for this pipeline:

• Flexiprep
• Alignment programs:
  • Bwa mem
  • Bwa aln
  • Bowtie version 1.1.1
  • Stampy
  • Gsnap
  • TopHat
  • Star
  • Star-2pass
• Picard tool suite

Configuration and flags

For technical reasons, single sample pipelines, such as this mapping pipeline do not take a sample config. Input files are in stead given on the command line as a flag.

Command line flags for the mapping pipeline are:

Flag (short)	Flag (long)	Type	Function
-R1	--input_r1	Path (required)	Path to input fastq file
-R2	--input_r2	Path (optional)	Path to second read pair fastq file.
-sample	--sampleid	String (required)	Name of sample
-library	--libid	String (required)	Name of library

If -R2 is given, the pipeline will assume a paired-end setup.

Sample input extensions

It is a good idea to check the format of your input files before starting any pipeline. Since the pipeline expects a specific format based on the file extensions. So for example if one inputs files with a fastq | fq extension the pipeline expects an unzipped fastq file. When the extension ends with fastq.gz | fq.gz the pipeline expects a bgzipped or gzipped fastq file.

Config

All other values should be provided in the config. Specific config values towards the mapping pipeline are:

Name	Type	Function
output_dir	Path (required)	directory for output files
reference_fasta	Path (required)	Path to indexed fasta file to be used as reference
aligner	String (optional)	Which aligner to use. Defaults to bwa. Choose from [bwa, bwa-aln, bowtie, gsnap, tophat, stampy, star, star-2pass]
skip_flexiprep	Boolean (optional)	Whether to skip the flexiprep QC step (default = False)
skip_markduplicates	Boolean (optional)	Whether to skip the Picard Markduplicates step (default = False)
skip_metrics	Boolean (optional)	Whether to skip the metrics gathering step (default = False)
platform	String (optional)	Read group Platform (defaults to illumina)
platform_unit	String (optional)	Read group platform unit
readgroup_sequencing_center	String (optional)	Read group sequencing center
readgroup_description	String (optional)	Read group description
predicted_insertsize	Integer (optional)	Read group predicted insert size

It is possible to provide any config value as a command line argument as well, using the -cv flag. E.g. -cv reference=<path/to/reference> would set value reference.

Example

Note that one should first create the appropriate settings config. Any supplied sample config will be ignored.

Example config

Minimal

{
"reference_fasta": "<path/to/reference">,
"output_dir": "<path/to/output/dir">
}

With options

{
"reference_fasta": "<path/to/reference">,
"aligner": "bwa",
"skip_metrics": true,
"platform": "our_platform",
"platform_unit":  "our_unit",
"readgroup_sequencing_center": "our_center",
"readgroup_description": "our_description",
"predicted_insertsize": 300,
"output_dir": "<path/to/output/dir">
}

Running the pipeline

For the help menu:

biopet pipeline mapping -h

Arguments for Mapping:
 -R1,--input_r1 <input_r1>             R1 fastq file
 -R2,--input_r2 <input_r2>             R2 fastq file
 -sample,--sampleid <sampleid>         Sample ID
 -library,--libid <libid>              Library ID
 -config,--config_file <config_file>   JSON / YAML config file(s)
 -cv,--config_value <config_value>     Config values, value should be formatted like 'key=value' or
                                       'path:path:key=value'
 -DSC,--disablescatter                 Disable all scatters

To run the pipeline:

biopet pipeline mapping -run --config mySettings.json \
-R1 myReads1.fastq -R2 myReads2.fastq

Note that removing -R2 causes the pipeline to assume single end .fastq files.

To perform a dry run simply remove -run from the commandline call.

---

Result files

├── OutDir
    ├── <samplename>-lib_1.dedup.bai
    ├── <samplename>-lib_1.dedup.bam
    ├── <samplename>-lib_1.dedup.metrics
    ├── flexiprep
    ├── metrics
    └── report

Getting Help

If you have any questions on running Mapping, suggestions on how to improve the overall flow, or requests for your favorite aligner to be added, feel free to post an issue to our issue tracker at GitHub. Or contact us directly via: SASC email