NeatSeq-Flow Cheat-Sheet

Author: Menachem Sklarz

Input Files

Sample file

Passed to NeatSeq-Flow with the -s argument.

Includes four sections:


A title for the project:

Title        Project_title

Project file information

Two tab-separated columns:

  1. File type
  2. File path
#Type        Path
Nucleotide   /path/to/genome.fasta

Samples file information

Three tab-separated columns:

  1. Sample ID
  2. File type
  3. File path

Additional columns will be ignored:

#SampleID    Type    Path    lane
Sample1      Forward /path/to/Sample1_F1.fastq.gz    1
Sample1      Forward /path/to/Sample1_F2.fastq.gz    2
Sample1      Reverse /path/to/Sample1_R1.fastq.gz    1
Sample1      Reverse /path/to/Sample1_R2.fastq.gz    2


Define ChIP and Control (‘input’) pairs:

Sample_Control      anti_sample1:input_sample1
Sample_Control      anti_sample2:input_sample2

Parameter file

Passed to NeatSeq-Flow with the -p argument.

YAML-formatted file with the following three sections.


The Vars section is recommended but not compulsory.

Global parameters section

Global parameters
Parameter Description
Executor SGE, Local or SLURM. (Default: SGE)
Qsub_q The cluster queue (or partition) to use. Default value for qsub –q parameter. Required
Qsub_nodes Default nodes on which to execute jobs (Default: All nodes in queue)
Qsub_opts Other parameters to pass to qsub
Qsub_path The full path to qsub. Obtain by running which qsub (default: qsub is in path)
Default_wait Default: 10. Leave as is
module_path List of paths to repositories of additional modules. (Must be a python directory, containing
Path to a file, defining parameters for limiting number of concurrent jobs, with the following line::
limit=1000 sleep=60
conda path and env, defining the path to the environment you want to use and its name (see here).
setenv Setting in global parameters is equivalent to setting setenv in all steps (see section Additional parameters.


The default executor is SGE. For SLURM, sbatch is used instead of qsub, e.g. Qsub_nodes defines the nodes to be used by sbatch.


If NeatSeq-Flow is executed from within a conda environment with both NeatSeq-Flow and it’s modules installed, module_path will automatically include the modules repo. If not, you will have to give the path to the location where the modules were installed.

Vars section

Replacements to be made in the parameter file. In YAML format. Referred to in other sections by the dot-notification.


    bwa:        /path/to/bwa
    samtools:   /path/to/samtools
  genome:       /path/to/genomeDir

In parameter section:

This… Becomes this…
{Vars.paths.bwa} /path/to/bwa
{Vars.paths.samtools} /path/to/samtools
{Vars.genome} /path/to/genomeDir

Step-wise parameters

A series of YAML blocks, one per workflow step to perform. Each block takes the following form:

  module:         fastqc_html
  base:           trim_gal
  script_path:    {Vars.paths.fastqc}

Types of step parameters:

Required parameters
Required parameters
Parameter Description
module The name of the module of which this step is an instance.
base The name of the step(s) on which the current step is based (not required for the Import step, which is always first and single)
script_path The full path to the script executed by this step.
Cluster parameters

Passed in a qsub_params block.

Cluster parameters
Parameter Description
node A node or YAML list of nodes on which to run the step scripts (overrides global parameter Qsub_nodes)
queue or -q Will limit the execution of the step’s scripts to this queue (overrides global parameter Qsub_q)
-pe Will set the -pe parameter for all scripts for this module (see SGE qsub manual).
-XXX: YYY Set the value of qsub parameter -XXX to YYY. This is a way to define other SGE parameters for all step scripts.
Additional parameters
Additional parameters
Parameter Description
tag All instances downstream to the tagged instance will have the same tag. All steps with the same tag can be executed with one master script
intermediate Will add a line to scripts/ for deleting the results of this step
setenv Set various environment variables for the duration of script execution. A string with format ENV="value for env1" ENV2="new value for env2"
precode Additional code to be added before the actual script. Rarely used
scope Use sample- or project-wise files. Check per-module documentation for whether and how this parameter is defined
sample_list Limit this step to a subset of the samples. See section Sample list.
conda Is used to define step specific conda parameters. The syntax is the same as for the global conda definition (see here).
arg_separator Set the delimiter between program argument and value, e.g. ‘=’ (Default: ‘ ‘)
local Use a local directory for intermediate files before copying results to final destination in data dir.
Redirected parameters

Parameters to be redirected to the actual program executed by the step.

Redirected parameters are specified within a redirects: block. The parameter name must include the - or -- required by the program defined in script_path.

Sample list

The sample list enables limiting the instance scripts to a subset of the samples. It can be expressed in two ways:

  1. A YAML list or a comma-separated list of sample names:

    sample_list: [sample1, sample2]
  2. By levels of a category (see section Mapping file):

        category:  Category1
        levels:     [level1,level2]

For using all but a subset of samples, use exclude_sample_list instead of sample_list.

Mapping file

Passed to NeatSeq-Flow with --mapping.

A tab-separated table with at least two columns:

  1. Sample ID
  2. First category name
  3. Additional categories…


#SampleID    Category1       Category2
Sample1      A       C
Sample2      A       D
Sample3      B       C
Sample4      B       D

Flow control


Basic mode

NeatSeq-Flow will attempt to guess all the parameters it requires.


    module:         Import

Advanced mode

Define source and target slots and how to concatenate the files. Attempts to guess information left out by the user.

Import parameters
Parameter Description
src source slot.
trg target slot
ext concatenated file extension.
scope the scope of the file
script_path the code to use for merging, or one of the following values:
pipe a command through which to pipe the file before storing.
Special values
Value Description
..guess.. Guess (script_path, trg and ext)
..import.. Do not copy the file, just import it into its slot (only if one file defined for src).
..skip.. Do not import the file type.


    module:         Import
    src:            [Forward,    Reverse, Nucl]
    trg:            [fastq.F,    fastq.R, fasta.nucl]
    script_path:    [..import.., cat,     'curl -L']
    ext:            [null,       null,    txt]
    scope:          [sample,     sample,  project]


Import raw data files into the data/ directory.

manage_types values
Value Possible values Description
operation add | del | mv | cp The operation to perform on the file type.
scope project|sample The scope on which to perform the operation. (For ‘mv’ and ‘cp’ this is the source scope)
type   The file type on which to perform the operation. (For ‘mv’ and ‘cp’ this is the source type)
scope_trg project|sample The destination scope for ‘mv’ and ‘cp’ operations
type_trg   The destination type for ‘mv’ and ‘cp’ operations.
Path   For add operation, the value to insert in the file type.


  module:   manage_types
  base:   trinity1
  scope:[project, sample, sample, project]
  operation: [mv,del,cp,add]
  type: [fasta.nucl, fasta.nucl, fastq.F, bam]
  type_trg:   [transcripts.nucl, None ,fastq.main, None]
  scope_trg:   sample
  path:   [None, None, None, /path/to/mapping.bam]


Used for concatenating tables from samples into one project table, or for concatenating tables from sample sub-samples, according to a mapping file. Any text file can be merged in this way.

merge_table parameters
Parameter Description
header The number of header lines the files contain.
add_filename Set to append the source filename to each line in the resulting file.
ext The extension to use in the resulting file. If not specified, uses merged file exts.
scope project or group, if group, you must also specify category.


    module:  merge_table
    base:      merge1
    type:  [blast.prot,fasta.nucl]
    header: 0
    ext:  [out,fna]

Reserved words

When writing new modules, the following words are conserved and should not be used for as parameters:

  • module
  • base
  • script_path
  • setenv
  • redirect
  • qsub_params
  • tag
  • conda
  • precode