Analyzing and allele-calling of MPS FASTQ files. This software is part of the DNASeqEx project.
These instructions will get you a copy of the project up and running on your local machine for development and testing purposes.
As this project is written Ruby, you obviously need to have a Ruby interpreter available. Please see Install Ruby to install Ruby.
ruby -v
First we need to install a ruby package manager called bundler. Please see Install Bundler to do so.
That will give us the needed bundle command.
bundle -v
At the main dirctory of this project
bundle install
will install all dependencies automatically.
We want to analyse only the STRs of an Illumina FGx HSC FASTQ file pair (read R1 and R2 are in two different files) in the example folder of our project:
ls -l example
-rwx------ 1 me staff 9,8M 8 Sep 2016 D701-D501_S1_L001_R1_001.fastq.gz
-rwx------ 1 me staff 843K 8 Sep 2016 D701-D501_S1_L001_R2_001.fastq.gz
To call the actual program we enter
./wintermute.rb --only forenseq --min-reads 10 --min-call-ratio 0.1 --r2-reverse-complement --no-n-trimming example/D701-D501_S1_L001_R1_001.fastq.gz
The resulting example/D501_S1_L001_CALL.xlsx is out output file. Please note that --r2-reverse-complement is neccessary for Illumina FGx runs only. In reasearch mode R2 reads are already in reverse-complent order.
Usage: ./wintermute.rb [options] <R1 FILE>
--[no-]calling [Do]/[Don't do] STR/SNP calling (default: true)
--[no-]survey [Do]/[Don't] summarize all assigned sequences into one directory/file (default: false)
--[no-]statistics [Do]/[Don't] output a separate statistics file (default: false)
-v, --[no-]verbose Run verbosely (default: false)
-c, --config <filename> Configuarion file for marker and target definitions (default: "./config/generic_grch38.json")
-k, --kits <filename> Kit configuarion file to name markers included in kits (default: "./config/kits.json")
-f, --[no-]force Overwrite result file(s) (default: false)
-o, --output-calling FILE STR/SNP calls output filename
-r, --references FILE Assign sequences to references stored in FILE
--[no-]dynamic-q Determine minimal Q-value dynamically based on R1/R2 (default: true)
--[no-]n-trimming Trim at first N (default: true)
--append Append results to existing file (requires -o)
--r1-reverse-complement Reverse complement the R1 sequence prior use (default: false)
--r2-reverse-complement Reverse complement the R1 sequence prior use (default: false)
--[no-]r2 Do/Don't automatically determine and load R2 (default: false)
--ignore-q Don't give a shit on the q-values (default: false)
--[no-]adapter-trimming Do/Don't trim adapter sequences (default: true)
--[no-]primer-trimming Do/Don't trim primer sequences (default: true)
--require-adapter Filter sequences that doesn't meet adapter requirements (specified in config file) (default: false)
--only [x,y,z] Process [x,y,z] targets/markers only. Kit names act as placeholders for the provided markers given at config/kits.json. (default: all)
--max-distance-of-forward-sequence-from-start N
Maximal distance from forward-sequence to read begin ('-1' disables this setting, default: -1)
--[no-]reversify-targets [Not] Reverse the configured targets to match reverse-complemnt as well (default: true)
--no-match-forward Sequence matching (STR/SNP) is performed only by matchng with reverse primer (default: true)
--no-match-reverse Sequence matching (STR/SNP) is performed only by matchng with forward primer (default: true)
--max-n N Maximal number of N within matching primers and R1/R2 first 20 bases (default: 3)
--min-q N Minimal Q-value for R1/R2 (overrides dynamic)
--min-reads N Minimal reads to consider a sequence (default: 10)
--min-reads-ratio N Minimal ratio of reads (relative to summarized target reads) to consider a sequence (default: 0.01)
--min-variant-reads N Minimal reads to consider a variant
--min-variant-reads-ratio N Minimal ratio of reads (relative to summarized target reads with same length) to consider a variant (default: 0.05)
--min-call-ratio N Minimal ratio of reads (relative to summarized target reads) to call a sequence.
The file config/generic_grch38.json contains almost al forensic relevant STR targets/markes. Please modify config/kits.json to fit your needs.
- highline - A higher level command-line oriented interface.
- rubyXL - Ruby lib for reading/writing/modifying .xlsx and .xlsm files
- parallel - Ruby: parallel processing made simple and fast
- pry - An IRB alternative and runtime developer console
- levenshtein-ffi - Fast string edit distance computation, using the Damerau-Levenshtein algorithm
Please read CONTRIBUTING.md for details on our code of conduct, and the process for submitting pull requests to us.
- Sascha Willuweit - Initial work
- Steffi Köcher - Testing
This project is licensed under the EUROPEAN UNION PUBLIC LICENCE v. 1.2 License - see the LICENSE.txt file for details
The DNASeqEx project has been funded with support from the European Commission (grant HOME/2014/ISFP/AG/LAWX/4000007135 under the Internal Security Funding Police programme of the European Commission-Directorate General Justice and Home Affairs).