Quick Start guide¶

First check that all requiered Dependancies are installed. If you wish to use AllMine on a cluster, contact your admnistrator for instalations.

Getting AllMine code¶

AllMine code is GitHub hosted. To get the source code using git CLI, use :

git clone https://github.com/tbersez/Allmine.git

Then, from within AllMine directory, build AllMine container using Singularity (admin rights needed) :

sudo singularity build AllMine singularity/AllMine_recipe.sr

Here, AllMine is ready to run.

Input files¶

To perform allele mining, AllMine needs :

Sequencing data (DNA or RNA)

A reference genome and annotation

A bed file with regions (or genes) of interest

Sequencing data¶

AllMine supports RNA and DNA sequencing, paired or single end, with a maximum read lenght of 350bp (you can submit longer reads but they will be trimmed to the maximum size). Sequencing data must be in fastq format and may be gzip compressed.

Sample sheet¶

Configuration maker for AllMine use a csv describing samples file as input. The csv file must have the flowing headers :

filename R1_ext R2_ext platform date(mm/dd/yy)

Here is an example:

"filename","R1_ext","R2_ext","platform","date(mm/dd/yy)"
"SRR1538456","_1.fastq.gz","_2.fastq.gz","illumina","03/03/03"
"SRR1538457","_1.fastq.gz","_2.fastq.gz","illumina","04/03/03"
"SRR1538484","_1.fastq.gz","_2.fastq.gz","illumina","05/03/03"

You can use a spread sheet editor to create this csv file !

Reference genome and annotation¶

Reference genome must be provided in one file, in fasta format. Annotation can be provided in gff or gtf format (recommended). When possible, we advice you to download the reference sequence and annotation from curated sources, such as Ensembl.

Bed file¶

Regions of interest must be specified using a bed file, here is an example :

NC_035163.1 25395963 25398308
NC_035168.1 31042453 31045884
NC_035169.1 25941228 25944616
NC_035175.1 3317633 3320503
NC_035177.1 20184932 20187543

Be sure that the first column (contigs) match with the ones used in your reference genome.

Making your work space ready¶

Place your reference genome and annotation in a common folder. That one must only contain those both files. Place your bed file with regions of interest in an other folder. AllMine outputs are created where your start the analysis. Make sure that you have enought space to store all outputs !

Configuration of an AllMine run¶

To configure an AllMine run use :

./csv_to_yaml.py path/to/sample_sheet.csv

Answer the questions the script is asking you to configure your run. Note : the bind path is the path from the root to your home folder.

Once done run :

./annovar_makebd.py

This script build the annotation database of Annovar. It need to done once for each new genome used.

Running AllMine¶

We recommend first to do a dry run using the following command. CORE_NUMBER must be replaced by the number of cores you wish to use.

snakemake -j CORE_NUMBER \
--cluster-config slurm_config.json \
--cluster "sbatch" -n

Check the output to ensure that your run is properly configured. If not, return to configuration step to correct errors. If yes, run :

snakemake -j CORE_NUMBER \
--cluster-config slurm_config.json \
--cluster "sbatch"

AllMine is now running. Depending on how much data you have submited and CORE_NUMBER, the analysis may take from few hours to a few days.