Falco: FastQC Alternative Code
This program is an emulation of the popular FastQC software to check large sequencing reads for common problems.
Installing falco
Installing through conda
If you use anaconda to manage your packages,
and the conda binary is in your path, you can install the most
recent release of falco by running
$ conda install -c bioconda falco
falco can be found inside the bin directory of your anaconda
installer.
Installing from source (code release)
Compilation from source can be done by downloading a falco release
from the releases
section above. Upon downloading the release (in .tar.gz or .zip
format), and inflating the downloaded file to a directory
(e.g. falco), move to the target directory the file was inflated
(e.g. cd falco). You should see a configure file in it. In this
directory, run
$ ./configure CXXFLAGS="-O3 -Wall"
$ make all
$ make install
if you wish to install the falco binaries on a specific directory, you can use
the --prefix argument when running ./configure, for instance:
$ ./configure CXXFLAGS="-O3 -Wall" --prefix=/path/to/installation/directory
The falco binary will be found in the bin directory inside the
specified prefix.
Installing from a cloned repository
We strongly recommend using falco through stable releases as
described above, as the latest commits might contain undocumented
bugs. For the more advanced users who wish to test the most recent
code, falco can be installed by first cloning the repository
$ git clone https://github.com/smithlabcode/falco.git
$ cd falco
Once inside the generated repsotory directory, run
$ make all
$ make install
This should create a bin directory inside the cloned repository
containing falco.
Required C++ dependencies
zlib is required to read gzip compressed FASTQ files. It is usually installed by default in most UNIX computers and is part of the htslib setup, but it can also be installed with standard package managers like apt, brew or conda.
On Ubuntu, zlib C++ libraries can be installed with apt:
$ sudo apt install zlib1g zlib1g-dev
Optional C++ dependencies
htslib is required to process bam files. If not provided, bam files will be treated as unrecognized file formats.
If htslib is installed, falco can be compiled with it by simply replacing the
configure command above with the --enable-hts flag:
$ ./configure CXXFLAGS="-O3 -Wall" --enable-hts
If falco was cloned from the repository, run the following commands
to allow BAM file reading:
$ make HAVE_HTSLIB=1 all
$ make HAVE_HTSLIB=1 install
If successfully compiled, falco can be used in BAM files the same way as it is
used with fastq and sam files.
Running falco
Run falco in with the following command, where the example.fq file
provided can be replaced with the path to any FASTQ file you want to run
falco
$ falco example.fq
This will generate three files in the same directory as the input fastq file:
-
fastqc_data.txtis a text file with a summary of the QC metrics -
fastqc_report.htmlis the visual HTML report showing plots of the QC metrics summarized in the text summary. -
summary.txt: A tab-separated file describing whether the pass/warn/fail result for each module. If multiple files are provided, only one summary file is generated, with one of the columns being the file name associated to each module result.
The full list of arguments and options can be seen by running falco
without any arguments, as well as falco -? or falco --help. This
will print the following list:
Usage: falco [OPTIONS] <seqfile1> <seqfile2> ...
Options:
-h, --help Print this help file and exit
-v, --version Print the version of the program and exit
-o, --outdir Create all output files in the specified
output directory. FALCO-SPECIFIC: If the
directory does not exists, the program will
create it. If this option is not set then
the output file for each sequence file is
created in the same directory as the
sequence file which was processed.
--casava [IGNORED BY FALCO] Files come from raw
casava output. Files in the same sample
group (differing only by the group number)
will be analysed as a set rather than
individually. Sequences with the filter flag
set in the header will be excluded from the
analysis. Files must have the same names
given to them by casava (including being
gzipped and ending with .gz) otherwise they
won't be grouped together correctly.
--nano [IGNORED BY FALCO] Files come from nanopore
sequences and are in fast5 format. In this
mode you can pass in directories to process
and the program will take in all fast5 files
within those directories and produce a
single output file from the sequences found
in all files.
--nofilter [IGNORED BY FALCO] If running with --casava
then don't remove read flagged by casava as
poor quality when performing the QC
analysis.
--extract [ALWAYS ON IN FALCO] If set then the zipped
output file will be uncompressed in the same
directory after it has been created. By
default this option will be set if fastqc is
run in non-interactive mode.
-j, --java [IGNORED BY FALCO] Provides the full path to
the java binary you want to use to launch
fastqc. If not supplied then java is assumed
to be in your path.
--noextract [IGNORED BY FALCO] Do not uncompress the
output file after creating it. You should
set this option if you do not wish to
uncompress the output when running in
non-interactive mode.
--nogroup Disable grouping of bases for reads >50bp.
All reports will show data for every base in
the read. WARNING: When using this option,
your plots may end up a ridiculous size. You
have been warned!
--min_length [NOT YET IMPLEMENTED IN FALCO] Sets an
artificial lower limit on the length of the
sequence to be shown in the report. As long
as you set this to a value greater or equal
to your longest read length then this will
be the sequence length used to create your
read groups. This can be useful for making
directly comaparable statistics from
datasets with somewhat variable read
lengths.
-f, --format Bypasses the normal sequence file format
detection and forces the program to use the
specified format. Valid formats are bam, sam,
bam_mapped, sam_mapped, fastq, fq, fastq.gz
or fq.gz.
-t, --threads [NOT YET IMPLEMENTED IN FALCO] Specifies the
number of files which can be processed
simultaneously. Each thread will be
allocated 250MB of memory so you shouldn't
run more threads than your available memory
will cope with, and not more than 6 threads
on a 32 bit machine [1]
-c, --contaminants Specifies a non-default file which contains
the list of contaminants to screen
overrepresented sequences against. The file
must contain sets of named contaminants in
the form name[tab]sequence. Lines prefixed
with a hash will be ignored. Default:
Configuration/contaminant_list.txt
-a, --adapters Specifies a non-default file which contains
the list of adapter sequences which will be
explicity searched against the library. The
file must contain sets of named adapters in
the form name[tab]sequence. Lines prefixed
with a hash will be ignored. Default:
Configuration/adapter_list.txt
-l, --limits Specifies a non-default file which contains
a set of criteria which will be used to
determine the warn/error limits for the
various modules. This file can also be used
to selectively remove some modules from the
output all together. The format needs to
mirror the default limits.txt file found in
the Configuration folder. Default:
Configuration/limits.txt
-k, --kmers [IGNORED BY FALCO AND ALWAYS SET TO 7]
Specifies the length of Kmer to look for in
the Kmer content module. Specified Kmer
length must be between 2 and 10. Default
length is 7 if not specified.
-q, --quiet Supress all progress messages on stdout and
only report errors.
-d, --dir [IGNORED: FALCO DOES NOT CREATE TMP FILES]
Selects a directory to be used for temporary
files written when generating report images.
Defaults to system temp directory if not
specified.
-s, -subsample [Falco only] makes falco faster (but
possibly less accurate) by only processing
reads that are multiple of this value (using
0-based indexing to number reads). [1]
-b, -bisulfite [Falco only] reads are whole genome
bisulfite sequencing, and more Ts and fewer
Cs are therefore expected and will be
accounted for in base content.
-r, -reverse-complement [Falco only] The input is a
reverse-complement. All modules will be
tested by swapping A/T and C/G
-skip-data [Falco only] Do not create FastQC data text
file.
-skip-report [Falco only] Do not create FastQC report
HTML file.
-skip-summary [Falco only] Do not create FastQC summary
file
-D, -data-filename [Falco only] Specify filename for FastQC
data output (TXT). If not specified, it will
be called fastq_data.txt in either the input
file's directory or the one specified in the
--output flag. Only available when running
falco with a single input.
-R, -report-filename [Falco only] Specify filename for FastQC
report output (HTML). If not specified, it
will be called fastq_report.html in either
the input file's directory or the one
specified in the --output flag. Only
available when running falco with a single
input.
-S, -summary-filename [Falco only] Specify filename for the short
summary output (TXT). If not specified, it
will be called fastq_report.html in either
the input file's directory or the one
specified in the --output flag. Only
available when running falco with a single
input.
-K, -add-call [Falco only] add the command call call to
FastQC data output and FastQC report HTML
(this may break the parse of fastqc_data.txt
in programs that are very strict about the
FastQC output format).
Help options:
-?, -help print this help message
-about print about message
PROGRAM: falco
A high throughput sequence QC analysis tool
Citing falco
If falco was helpful for your research, you can cite us as follows:
de Sena Brandine G and Smith AD. Falco: high-speed FastQC emulation for quality control of sequencing data. F1000Research 2021, 8:1874 (https://doi.org/10.12688/f1000research.21142.2)
Please do not cite this manuscript if you used FastQC directly and not falco!
Copyright and License Information
Copyright (C) 2019-2024 Guilherme de Sena Brandine and Andrew D. Smith
Authors: Guilherme de Sena Brandine and Andrew D. Smith
This is free software: you can redistribute it and/or modify it under the terms of the GNU General Public License as published by the Free Software Foundation, either version 3 of the License, or (at your option) any later version.
This software is distributed in the hope that it will be useful, but WITHOUT ANY WARRANTY; without even the implied warranty of MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the GNU General Public License for more details.