Metadata-Version: 1.1
Name: rmseq
Version: 0.0.45b0
Summary: RM-seq is a bioinformatics tool for for assessing resistance mutations from PE short-reads.
Home-page: UNKNOWN
Author: Romain Guerillot, Torsten Seemann, Mark B Schultz
Author-email: romain.guerillot@hotmail.fr
License: AGPL-3.0
Download-URL: https://github.com/rguerillot/RM-seq/zipball/master
Description-Content-Type: UNKNOWN
Description: RM-seq: Resistance Mutation SEQuencing
        ======================================
        
        Analysis bioinformatic pipeline for high-throughput identification and
        quantification of large repertoires of resistance conferring mutations.
        RM-seq is an amplicon-based, deep-sequencing technique using single
        molecule barcoding. We have adapted this method in order to identify and
        quantify mutations that confer resistance to a given antibiotic.
        
        RM-seq allows to both correct sequenced read errors generated during
        sequencing and to accurately quantify mutations by correcting PCR
        amplification bias generated during sequencing library preparation.
        During the first step of amplicon library preparation, a linear PCR
        (primer extension) with a primer comprising a tail with degenerated
        bases (all possible bases) introduce a unique barcode to DNA template
        molecules. Therefore a barcode is assigned not just to all the molecules
        from a certain sample (indexing), but to all molecules being amplified
        and sequenced. RM-seq pipeline use these barcodes to generate an
        error-corrected consensus sequence of the initial template variant.
        Counting the barcodes indroduced before exponential amplification by PCR
        of the template allows to accurately quantify each genetic variants from
        genomic DNA extracted from complex population of resistant clones (eg.
        pools of 10,000 resistant colonies selected by an antibiotic in vitro).
        
        A complete descrition of the RM-seq method will be available soon
        (article submitted)
        
        Is this the right tool for me?
        ------------------------------
        
        1. To be able to us this pipeline you need to have sequenced amplicon
           library with molecular barcodes.
        2. It only supports paired-end FASTQ reads (including .gz compressed
           fastq files).
        3. It needs paired reads that are overlapping.
        4. It needs a reference fasta sequences of the sequenced gene (DNA and
           protein sequence).
        5. It's written in Python3 and Perl.
        
        Installation
        ------------
        
        Install RM-seq pipeline
        ~~~~~~~~~~~~~~~~~~~~~~~
        
        ::
        
            pip3 install rmseq
        
        Dependencies
        ~~~~~~~~~~~~
        
        RM-seq has the following package dependencies: \* EMBOSS >= 6.6 for
        ``clustalo``, ``cons``, ``getorf``, ``diffseq`` \* clustal-omega >=
        1.2.1 \* bwa >= 0.7.15 \* samtools >= 1.3 \* bedtools >= 2.26.0 \* pear
        >= 0.9.10 \* cd-hit >= 4.7 \* trimmomatic >= 0.36 \* python modules:
        ``plumbum``, ``Biopython``
        
        If you are using the `OSX Brew <http://brew.sh/>`__ or
        `LinuxBrew <http://linuxbrew.sh/>`__ packaging system:
        
        ::
        
            brew tap homebrew/science
            brew tap tseemann/bioinformatics-linux
            brew install parallel; parallel --citation # please write will cite
            brew install bedtools
            brew install EMBOSS
            brew install clustal-omega
            brew install bwa
            brew install samtools
            brew install pear
            brew install cd-hit
            brew install trimmomatic
            pip3 install plumbum
            pip3 install biopython
        
        Quick start
        -----------
        
        Do
        
        ::
        
            rmseq
        
        Help
        ~~~~
        
        ::
        
            usage: rmseq [-h]  ...
        
            Run RM-seq pipeline.
        
            optional arguments:
              -h, --help  show this help message and exit
        
            Commands:
        
                run       Run the pipeline.
                version   Print version.
                check     Check pipeline dependencies
                test      Run the test data set.
        
        To check dependencies are installed
        ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
        
        ::
        
            rmseq check
        
        To run the test dataset
        ~~~~~~~~~~~~~~~~~~~~~~~
        
        ::
        
            rmseq test
        
        To run analysis pipeline, follow the steps in
        ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
        
        ::
        
            rmseq run -h
            usage: rmseq run [options]
        
            Run the pipeline
        
            positional arguments:
              R1                    Path to read pair 1
              R2                    Path to read pair 2
              refnuc                Reference gene that will be used for premapping
                                    filtering (fasta).
              refprot               Reference protein that will be use for annotating
                                    variants (fasta).
              outdir                Output directory.
        
            optional arguments:
              -h, --help            show this help message and exit
              -d, --debug_on        Switch on debug mode.
              -f, --force           Force overwite of existing.
              -b BARLEN, --barlen BARLEN
                                    Length of barcode (default 16)
              -m MINFREQ, --minfreq MINFREQ
                                    Minimum barcode frequency to keep (default 5)
              -c CPUS, --cpus CPUS  Number of CPUs to use (default 72)
              -r MINSIZE, --minsize MINSIZE
                                    Minimum ORF size in bp used when annotating variants
                                    (default 200)
              -w WSIZE, --wsize WSIZE
                                    Word-size option to pass to diffseq for comparison
                                    with reference sequence (default 5)
              -s SUBSAMPLE, --subsample SUBSAMPLE
                                    Only examine this many reads.
              -k, --keepfiles       Keep the intermediate files. Default is to remove
                                    intermediate files
        
        To check the version
        ~~~~~~~~~~~~~~~~~~~~
        
        ::
        
            rmseq version
        
        Outputs
        -------
        
        RM-seq produces a tap-separated output file called amplicons.effect with
        the following columns:
        
        +----------+------------+---------------+
        | Column   | Example    | Description   |
        +==========+============+===============+
        | barcode  | GACACAACTG | The sequence  |
        |          | AGATTA     | of the        |
        |          |            | barcode       |
        +----------+------------+---------------+
        | sample   | Rifampicin | The output    |
        |          | 1          | folder name   |
        +----------+------------+---------------+
        | aa\_muta | H481N      | The           |
        | tion     |            | annotation of |
        |          |            | the amino     |
        |          |            | acid change   |
        |          |            | (Histidine    |
        |          |            | residue 481   |
        |          |            | substituted   |
        |          |            | by            |
        |          |            | Asparagine)   |
        +----------+------------+---------------+
        | start    | 481        | start         |
        |          |            | coordinate of |
        |          |            | the mutation  |
        +----------+------------+---------------+
        | end      | 481        | end           |
        |          |            | coordinate of |
        |          |            | the mutation  |
        +----------+------------+---------------+
        | orf      | VRPPDKNNRF | the protein   |
        |          | VGLYCTLV.. | sequence of   |
        |          | .          | the consensus |
        |          |            | sequence      |
        +----------+------------+---------------+
        | dna      | GGTTAGACCA | The dna       |
        |          | CCCGATAA.. | sequence of   |
        |          | .          | the consensus |
        |          |            | sequence      |
        +----------+------------+---------------+
        
        The other files produced by RM-seq are:
        
        +-------------+---------------+
        | File name   | Description   |
        +=============+===============+
        | amplicons.n | Multifasta    |
        | uc          | file          |
        |             | containing    |
        |             | all the       |
        |             | consensus     |
        |             | nucleotide    |
        |             | sequence      |
        |             | (header of    |
        |             | sequence is   |
        |             | the barcode)  |
        +-------------+---------------+
        | amplicons.o | Multifasta    |
        | rf          | file          |
        |             | containing    |
        |             | all the       |
        |             | consensus     |
        |             | protein       |
        |             | sequence      |
        |             | (header of    |
        |             | sequence is   |
        |             | the barcode)  |
        +-------------+---------------+
        | amplicons.b | Table with    |
        | arcodes     | the count of  |
        |             | each barcode  |
        |             | sequence      |
        +-------------+---------------+
        | amplicons.c | Multifasta    |
        | dhit        | file          |
        |             | containing    |
        |             | all the       |
        |             | unique        |
        |             | consensus     |
        |             | nucleotide    |
        |             | sequence      |
        |             | (header of    |
        |             | sequence is   |
        |             | the barcode)  |
        +-------------+---------------+
        
        Issues
        ------
        
        Please report problems to the `Issues
        Page <https://github.com/rguerillot/RM-seq/issues>`__.
        
        Author
        ------
        
        Romain Guerillot \| Torsten Seemann \| Mark B Schultz (github: schultzm)
        
Keywords: resistance,mutation
Platform: UNKNOWN
Classifier: Development Status :: 3 - Alpha
Classifier: License :: OSI Approved :: GNU Affero General Public License v3 or later (AGPLv3+)
Classifier: Programming Language :: Python :: 3.5
Classifier: Topic :: Scientific/Engineering :: Bio-Informatics
Classifier: Topic :: Scientific/Engineering :: Medical Science Apps.
Classifier: Intended Audience :: Science/Research
