Overview

Teaching: 30 min
Exercises: 10 min

Questions

• How do I get data into Nextflow?

• How do I handle different types of input, e.g. files and parameters?

• How do I create a Nextflow channel?

• How can I use pattern matching to select input files?

• How do I change the way inputs are handled?

Objectives

• Understand how Nextflow manages data using channels.

• Understand the different types of Nextflow channels.

• Create a value and queue channel using channel factory methods.

• Select files as input based on a glob pattern.

• Edit channel factory arguments to alter how data is read in.

Channels

Earlier we mentioned that channels are the way in which Nextflow sends data around a workflow. Channels connect processes via their inputs and outputs. Channels can store multiple items, such as files (e.g., fastq files) or values.

Why use Channels?

Channels are how Nextflow handles file management, allowing complex tasks to be split up, run in parallel, and simplifies getting the right inputs to the right parts of the pipeline.

Channels are asynchronous, which means that outputs from a set of processes will not necessarily be produced in the same order as the corresponding inputs went in. However, the first element into a channel queue is the first out of the queue (First in - First out). This allows processes to run as soon as they receive input from a channel. Channels only send data in one direction, from a producer (a process/operator), to a consumer (another process/operator).

Channel types

Nextflow distinguishes between two different kinds of channels: queue channels and value channels.

Queue channel

Queue channels are a type of channel in which data is consumed (used up) to make input for a process/operator. Queue channels can be created in two ways:

1. As the outputs of a process.
2. Explicitly using channel factory methods such as Channel.of or Channel.fromPath.

DSL1

In Nextflow DSL1 queue channels can only be used once in a workflow, either connecting workflow input to process input, or process output to input for another process. In DSL2 we can use a queue channel multiple times.

Value channels

The second type of Nextflow channel is a value channel. A value channel is bound to a single value. A value channel can be used an unlimited number times since its content is not consumed. This is also useful for processes that need to reuse input from a channel, for example, a reference genome sequence file that is required by multiple steps within a process, or by more than one process.

Queue vs Value Channel.

What type of channel would you use to store the following?

1. Multiple values.
2. A list with one or more values.
3. A single value.

Solution

1. A queue channels is used to store multiple values.
2. A value channel is used to store a single value, this can be a list with multiple values.
3. A value channel is used to store a single value.

Creating Channels using Channel factories

Channel factories are used to explicitly create channels. In programming, factory methods (functions) are a programming design pattern used to create different types of objects (in this case, different types of channels). They are implemented for things that represent more generalised concepts, such as a Channel.

Channel factories are called using the Channel.<method> syntax, and return a specific instance of a Channel.

The value Channel factory

The value factory method is used to create a value channel. Values are put inside parentheses () to assign them to a channel.

For example:

ch1 = Channel.value( 'GRCh38' )
ch2 = Channel.value( ['chr1', 'chr2', 'chr3', 'chr4', 'chr5'] )
ch3 = Channel.value( ['chr1' : 248956422, 'chr2' : 242193529, 'chr3' : 198295559] )

1. Creates a value channel and binds a string to it.
2. Creates a value channel and binds a list object to it that will be emitted as a single item.
3. Creates a value channel and binds a map object to it that will be emitted as a single item.

The value method can only take 1 argument, however, this can be a single list or map containing several elements.

Reminder:

• A List object can be defined by placing the values in square brackets [] separated by a comma.
• A Map object is similar, but with key:value pairs separated by commas.

myList = [1776, -1, 33, 99, 0, 928734928763]
myMap = [ p1 : "start", q2 : "end" ]

Queue channel factory

Queue (consumable) channels can be created using the following channel factory methods.

• Channel.of
• Channel.fromList
• Channel.fromPath
• Channel.fromFilePairs
• Channel.fromSRA

The of Channel factory

When you want to create a channel containing multiple values you can use the channel factory Channel.of. Channel.of allows the creation of a queue channel with the values specified as arguments, separated by a ,.

chromosome_ch = Channel.of( 'chr1', 'chr3', 'chr5', 'chr7' )
chromosome_ch.view()

chr1
chr3
chr5
chr7

The first line in this example creates a variable chromosome_ch. chromosome_ch is a queue channel containing the four values specified as arguments in the of method. The view operator will print one line per item in a list. Therefore the view operator on the second line will print four lines, one for each element in the channel:

You can specify a range of numbers as a single argument using the Groovy range operator ... This creates each value in the range (including the start and end values) as a value in the channel. The Groovy range operator can also produce ranges of dates, letters, or time. More information on the range operator can be found here.

chromosome_ch = Channel.of(1..22, 'X', 'Y')
chromosome_ch.view()

Arguments passed to the of method can be of varying types e.g., combinations of numbers, strings, or objects. In the above examples we have examples of both string and number data types.

Channel.from

You may see the method Channel.from in older nextflow scripts. This performs a similar function but is now deprecated (no longer used), and so Channel.of should be used instead.

Create a value and Queue and view Channel contents

1. Create a Nextflow script file called channel.nf .
2. Create a Value channel ch_vl containing the String 'GRCh38'.
3. Create a Queue channel ch_qu containing the values 1 to 4.
4. Use .view() operator on the channel objects to view the contents of the channels.
5. Run the code using

   $ nextflow run channel.nf
   

Solution

ch_vl = Channel.value('GRCh38')
ch_qu = Channel.of(1,2,3,4)
ch_vl.view()
ch_qu.view()

 N E X T F L O W  ~  version 21.04.0
 Launching `channel.nf` [condescending_dalembert] - revision: c80908867b
 GRCh38
 1
 2
 3
 4

The fromList Channel factory

You can use the Channel.fromList method to create a queue channel from a list object.

aligner_list = ['salmon', 'kallisto']

aligner_ch = Channel.fromList(aligner_list)

aligner_ch.view()

This would produce two lines.

salmon
kallisto

Channel.fromList vs Channel.of

In the above example, the channel has two elements. If you has used the Channel.of(aligner_list) it would have contained only 1 element [salmon, kallisto] and any operator or process using the channel would run once.

Creating channels from a list

Write a Nextflow script that creates both a queue and value channel for the list

ids = ['ERR908507', 'ERR908506', 'ERR908505']

Then print the contents of the channels using the view operator. How many lines does the queue and value channel print?

Hint: Use the fromList() and value() Channel factory methods.

Solution

ids = ['ERR908507', 'ERR908506', 'ERR908505']

queue_ch = Channel.fromList(ids)
value_ch = Channel.value(ids)
queue_ch.view()
value_ch.view()

N E X T F L O W  ~  version 21.04.0
Launching `channel_fromList.nf` [wise_hodgkin] - revision: 22d76be151
[ERR908507, ERR908506, ERR908505]
ERR908507
ERR908506
ERR908505

The queue channel queue_ch will print three lines.

The value channel value_ch will print one line.

The fromPath Channel factory

The previous channel factory methods dealt with sending general values in a channel. A special channel factory method fromPath is used when wanting to pass files.

The fromPath factory method creates a queue channel containing one or more files matching a file path.

The file path (written as a quoted string) can be the location of a single file or a “glob pattern” that matches multiple files or directories.

The file path can be a relative path (path to the file from the current directory), or an absolute path (path to the file from the system root directory - starts with /).

The script below creates a queue channel with a single file as its content.

read_ch = Channel.fromPath( 'data/yeast/reads/ref1_2.fq.gz' )
read_ch.view()

data/yeast/reads/ref1_2.fq.gz

You can also use glob syntax to specify pattern-matching behaviour for files. A glob pattern is specified as a string and is matched against directory or file names.

• An asterisk, *, matches any number of characters (including none).
• Two asterisks, **, works like * but will also search sub directories. This syntax is generally used for matching complete paths.
• Braces {} specify a collection of subpatterns. For example: {bam,bai} matches “bam” or “bai”

For example the script below uses the *.fq.gz pattern to create a queue channel that contains as many items as there are files with .fq.gz extension in the data/yeast/reads folder.

read_ch = Channel.fromPath( 'data/yeast/reads/*.fq.gz' )
read_ch.view()

data/yeast/reads/ref1_2.fq.gz
data/yeast/reads/etoh60_3_2.fq.gz
data/yeast/reads/temp33_1_2.fq.gz
data/yeast/reads/temp33_2_1.fq.gz
data/yeast/reads/ref2_1.fq.gz
data/yeast/reads/temp33_3_1.fq.gz
[..truncated..]

Note The pattern must contain at least a star wildcard character.

You can change the behaviour of Channel.fromPath method by changing its options. A list of .fromPath options is shown below.

Available fromPath options:

We can change the default options for the fromPath method to give an error if the file doesn’t exist using the checkIfExists parameter. In Nextflow, method parameters are separated by a , and parameter values specified with a colon :.

If we execute a Nextflow script with the contents below, it will run and not produce an output. This is likely not what we want.

read_ch = Channel.fromPath( 'data/chicken/reads/*.fq.gz' )
read_ch.view()

Add the argument checkIfExists with the value true.

read_ch = Channel.fromPath( 'data/chicken/reads/*.fq.gz', checkIfExists: true )
read_ch.view()

This will give an error as there is no data/chicken directory.

N E X T F L O W  ~  version 20.10.0
Launching `hello.nf` [intergalactic_mcclintock] - revision: d2c138894b
No files match pattern `*.fq.gz` at path: data/chicken/reads/

Using Channel.fromPath

1. Create a Nextflow script channel_fromPath.nf
2. Use the Channel.fromPath method to create a channel containing all files in the data/yeast/ directory, including the subdirectories.
3. Add the parameter to include any hidden files.
4. Then print all file names using the view operator.

Hint: You need two asterisks, i.e. **, to search subdirectories.

Solution

all_files_ch = Channel.fromPath('data/yeast/**', hidden: true)
all_files_ch.view()

N E X T F L O W  ~  version 21.04.0
Launching `nf-training/scripts/channels/channel_fromPath.nf` [reverent_mclean] - revision: cf02269bcb
data/yeast/samples.csv
data/yeast/.hidden_file.txt
data/yeast/bams/ref1.bam.bai
data/yeast/bams/ref3.bam
data/yeast/bams/etoh60_3.
[..truncated..]

The fromFilePairs Channel factory

We have seen how to process files individually using fromPath. In Bioinformatics we often want to process files in pairs or larger groups, such as read pairs in sequencing.

For example is the data/yeast/reads directory we have nine groups of read pairs.

Nextflow provides a convenient Channel factory method for this common bioinformatics use case. The fromFilePairs method creates a queue channel containing a tuple for every set of files matching a specific glob pattern (e.g., /path/to/*_{1,2}.fq.gz).

A tuple is a grouping of data, represented as a Groovy List.

1. The first element of the tuple emitted from fromFilePairs is a string based on the shared part of the filenames (i.e., the * part of the glob pattern).
2. The second element is the list of files matching the remaining part of the glob pattern (i.e., the <string>_{1,2}.fq.gz pattern). This will include any files ending _1.fq.gz or _2.fq.gz.

read_pair_ch = Channel.fromFilePairs('data/yeast/reads/*_{1,2}.fq.gz')
read_pair_ch.view()

[etoh60_3, [data/yeast/reads/etoh60_3_1.fq.gz, data/yeast/reads/etoh60_3_2.fq.gz]]
[temp33_1, [data/yeast/reads/temp33_1_1.fq.gz, data/yeast/reads/temp33_1_2.fq.gz]]
[ref1, [data/yeast/reads/ref1_1.fq.gz, data/yeast/reads/ref1_2.fq.gz]]
[ref2, [data/yeast/reads/ref2_1.fq.gz, data/yeast/reads/ref2_2.fq.gz]]
[temp33_2, [data/yeast/reads/temp33_2_1.fq.gz, data/yeast/reads/temp33_2_2.fq.gz]]
[ref3, [data/yeast/reads/ref3_1.fq.gz, data/yeast/reads/ref3_2.fq.gz]]
[temp33_3, [data/yeast/reads/temp33_3_1.fq.gz, data/yeast/reads/temp33_3_2.fq.gz]]
[etoh60_1, [data/yeast/reads/etoh60_1_1.fq.gz, data/yeast/reads/etoh60_1_2.fq.gz]]
[etoh60_2, [data/yeast/reads/etoh60_2_1.fq.gz, data/yeast/reads/etoh60_2_2.fq.gz]]

This will produce a queue channel, read_pair_ch , containing nine elements.

Each element is a tuple that has;

1. string value (the file prefix matched, e.g temp33_1)
2. and a list with the two files e,g. [data/yeast/reads/temp33_1_1.fq.gz, data/yeast/reads/temp33_1_2.fq.gz] .

The asterisk character *, matches any number of characters (including none), and the {} braces specify a collection of subpatterns. Therefore the *_{1,2}.fq.gz pattern matches any file name ending in _1.fq.gz or _2.fq.gz .

What if you want to capture more than a pair?

If you want to capture more than two files for a pattern you will need to change the default size argument (the default value is 2) to the number of expected matching files.

For example in the directory data/yeast/reads there are six files with the prefix ref. If we want to group (create a tuple) for all of these files we could write;

read_group_ch = Channel.fromFilePairs('data/yeast/reads/ref{1,2,3}*',size:6)
read_group_ch.view()

The code above will create a queue channel containing one element. The element is a tuple of which contains a string value, that is the pattern ref, and a list of six files matching the pattern.

[ref, [data/yeast/reads/ref1_1.fq.gz, data/yeast/reads/ref1_2.fq.gz, data/yeast/reads/ref2_1.fq.gz, data/yeast/reads/ref2_2.fq.gz, data/yeast/reads/ref3_1.fq.gz, data/yeast/reads/ref3_2.fq.gz]]

See more information about the channel factory fromFilePairs here

More complex patterns

If you need to match more complex patterns you should create a sample sheet specifying the files and create a channel from that. This will be covered in the operator episode.

Create a channel containing groups of files

1. Create a Nextflow script file channel_fromFilePairs.nf .
2. Use the fromFilePairs method to create a channel containing three tuples. Each tuple will contain the pairs of fastq reads for the three temp33 samples in the data/yeast/reads directory

Solution

pairs_ch = Channel.fromFilePairs('data/yeast/reads/temp33*_{1,2}.fq.gz')
pairs_ch.view()

N E X T F L O W  ~  version 21.04.0
Launching `channels.nf` [stupefied_lumiere] - revision: a3741edde2
[temp33_1, [data/yeast/reads/temp33_1_1.fq.gz, data/yeast/reads/temp33_1_2.fq.gz]]
[temp33_3, [data/yeast/reads/temp33_3_1.fq.gz, data/yeast/reads/temp33_3_2.fq.gz]]
[temp33_2, [data/yeast/reads/temp33_2_1.fq.gz, data/yeast/reads/temp33_2_2.fq.gz]]

The fromSRA Channel factory

Another useful factory method is fromSRA. The fromSRA method makes it possible to query the NCBI SRA archive and returns a queue channel emitting the FASTQ files matching the specified selection criteria.

The queries can be project IDs or accession numbers supported by the NCBI ESearch API.

If you want to use this functionality, you will need an NCBI API KEY, and to set the environment variable NCBI_API_KEY to its value.

sra_ch =Channel.fromSRA('SRP043510')
sra_ch.view()

This will print a tuple for every fastq file associated with that SRA project accession.

[SRR1448794, ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR144/004/SRR1448794/SRR1448794.fastq.gz]
[SRR1448795, ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR144/005/SRR1448795/SRR1448795.fastq.gz]
[SRR1448792, ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR144/002/SRR1448792/SRR1448792.fastq.gz]
[SRR1448793, ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR144/003/SRR1448793/SRR1448793.fastq.gz]
[SRR1910483, ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR191/003/SRR1910483/SRR1910483.fastq.gz]
[SRR1910482, ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR191/002/SRR1910482/SRR1910482.fastq.gz]
(remaining omitted)

Multiple accession IDs can be specified using a list object:

ids = ['ERR908507', 'ERR908506', 'ERR908505']
sra_ch = Channel.fromSRA(ids)
sra_ch.view()

[ERR908507, [ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR908/ERR908507/ERR908507_1.fastq.gz, ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR908/ERR908507/ERR908507_2.fastq.gz]]
[ERR908506, [ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR908/ERR908506/ERR908506_1.fastq.gz, ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR908/ERR908506/ERR908506_2.fastq.gz]]
[ERR908505, [ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR908/ERR908505/ERR908505_1.fastq.gz, ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR908/ERR908505/ERR908505_2.fastq.gz]]

Read pairs from SRA

Read pairs are implicitly managed, and are returned as a list of files.

Key Points

• Channels must be used to import data into Nextflow.

• Nextflow has two different kinds of channels: queue channels and value channels.

• Data in value channels can be used multiple times in workflow.

• Data in queue channels are consumed when they are used by a process or an operator.

• Channel factory methods, such as Channel.of, are used to create channels.

• Channel factory methods have optional parameters e.g., checkIfExists, that can be used to alter the creation and behaviour of a channel.