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cld.pl
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#!/usr/bin/perl
###################################################################################################################################################################################################
# initialize geneneral cld settings and import perl dependencies
###################################################################################################################################################################################################
#use strict;
#use warnings FATAL => 'all';
use Bio::DB::Fasta;
use Bio::SeqIO;
use Bio::Tools::GFF;
use Scalar::Util qw(looks_like_number);
use Bio::SeqFeature::Generic; #important package to handle sequence formats and objects
use Bio::Location::Split; #library to make splitted location objects
use Set::IntervalTree; #library providing methods for using interval trees
use JSON::XS qw(encode_json decode_json); #library to encode perl objects in a file
use File::Slurp qw(read_file write_file); #library to write perl objects to a file
use List::MoreUtils qw{
any all none notall true false
firstidx first_index lastidx last_index
insert_after insert_after_string
apply indexes
after after_incl before before_incl
firstval first_value lastval last_value
each_array each_arrayref
pairwise natatime
mesh zip uniq distinct minmax part}; #some math and list utilities, later needed to partition a list of entries
use List::Util qw(sum);
use Archive::Zip;
use Parallel::ForkManager; #important package to enable mutlithreading of the script
use Cwd;
use IPC::Cmd qw[can_run run];
use Getopt::Long qw(:config pass_through);
use File::Grep qw( fgrep fmap fdo );
use Text::Wrap;
use Unix::Processors;
require Tk; #loads the Tk library, neccessary for producing a simple Tk GUI
Tk->import;
require Tk::PathEntry; #loads the interactive pathentry widget
require Tk::Dialog; #loads the Tk Fileopener dialog widget
require Tk::Dressing;
require Tk::BrowseEntry;
require Tk::Optionmenu;
require Tk::Widget;
require Tk::Frame;
require Tk::Entry;
require Tk::Label;
require Tk::Button;
require Tk::Scrollbar;
require Tk::Checkbutton;
require Tk::MainWindow;
require Tk::NoteBook;
require Tk::Text::SuperText;
#open(my $debug_log, ">", "debug_log.txt") or die $!;
my $procs = new Unix::Processors;
my $max_parallel= my $parallel_number =$procs->max_online;
my $aligner_path="";
if(-d $ENV{PAR_TEMP}."/inc/"){
$aligner_path=$ENV{PAR_TEMP}."/inc/bowtie_progs/";
}
$| = 1;
my ($script_name,$script_version,$script_date,$script_years) = ('cld','1.3.1','2015-11-24','2013-2015');
###################################################################################################################################################################################################
# initialize all GUI and command line variables
###################################################################################################################################################################################################
my(
%parameter_presets,
%something,
$annotation_options_frame,
$annotation_options_page,
$chk_CDS_only,
$chk_CpG_exclusive,
$chk_draw_html_report,
$annonymous_funct,
$chk_exclude_overlapping_genes,
$chk_exon_exclusive,
$chk_gene_exclusive,
$chk_ignore_intergenic,
$chk_ignore_missing_id,
$chk_match_info,
$chk_out_gff,
$chk_purpose_exclusive,
$chk_retrieve_recomb_matrix,
$chk_sec_off_target,
$chosen_kit,
$databasepath_entry,
$general_options_frame,
$general_options_page,
$io_options_frame,
$io_options_page,
$key,
$kit_frame,
$kit_list,
$kit_page,
$lab_crisprai,
$lab_databasepath,
$lab_kit_list,
$lab_knockout,
$lab_paired,
$lab_ref_organism,
$lab_specific_exon,
$lab_specific_transcript,
$lab_tagging,
$list_lab,
$mw,
$nb,
$off_target_options_page,
$offtarget_options_frame,
$opt_bowtie_mode,
$opt_bowtie_version,
$opt_data_type,
$gene_list_entry,
$opt_kind,
$opt_offtargetdb,
$opt_PAM,
$opt_preceding,
$opt_purpose,
$param,
$ref_organism_entry,
$scl_crispra_downstream,
$scl_crispra_upstream,
$scl_crispri_downstream,
$scl_crispri_upstream,
$scl_downstream_window,
$scl_edit_distance_allowed,
$scl_left_homology,
$scl_max_A,
$scl_max_C,
$scl_max_G,
$scl_max_length,
$scl_max_per_exon,
$scl_max_T,
$scl_maxspacerlength,
$scl_min_A,
$scl_min_C,
$scl_min_G,
$scl_min_length,
$scl_min_T,
$scl_minspacerlength,
$scl_number_of_CDS,
$scl_off_targets_allowed,
$scl_right_homology,
$scl_unspecific_leading_bases,
$scl_upstream_window,
$specific_exon_entry,
$specific_options_frame,
$specific_options_page,
$specific_options_placeholder_frame,
$specific_transcripts_entry,
$Start_but,
$temp_file,
$theme,
$tk_dressing,
$trigger_frame,
$trigger_page,
$higher_order_functions,
$lower_order_functions,
$make_database,
$target_ident,
$end_to_end,
$gene_list,
$database_download_but,
$database_creation_but,
$opt_rsync,
$opt_precalc,
$lab_gene_entry,
@string,
$isok,
$lab_output_dir,
$output_dir_entry ,
$funct_results
);
###################################################################################################################################################################################################
# read in command line options
###################################################################################################################################################################################################
GetOptions(
'task=s' => \$something{"task"},
'output-dir=s' => \$something{"working_path"},
'parameter-file=s' => \$something{"param_file_name"},
'gene-list=s' => \$something{"gene_list_file_name"},
'cov=i' => \$something{"coverage"},
'lib-size=i' => \$something{"total_lib_size"},
'library_name=s' => \$something{"library_name"},
'5-prime=s' => \$something{"5_adapt"},
'3-prime=s' => \$something{"3_adapt"},
'cor-5-prime=s' => \$something{"correct_5_prime_G"},
'input-folder=s' => \$something{"input_folder"},
'organism=s' => \$something{"organism_db"},
'rsync-link=s' => \$something{"rsync_link"},
'version' => \$something{"version"},
'help' => \$something{"help"},
'GUI' => \$something{"GUI"},
'spread-over-transcripts=s'=> \$something{"cover_many_transcripts"},
'scoring-module=s'=>\$something{"scoring_module"}
);
###################################################################################################################################################################################################
# define help and version strings
###################################################################################################################################################################################################
$something{"version_string"} = "$script_name, version $script_version, $script_date\nAuthor $script_years Florian Heigwer\n";
$something{"help_string"} = qq{Usage: cld --task=end_to_end [options=value] ...
Options:
--task=<task option>
make_database to provide an cld ready data base.
--organism=<string> Specify an organism to build the database for.
it must be one of the organisms available in ENSEMBLs ftp repository.
And in the same format as its ENSEMBL ftp directoy name.
E.g.: drosophila_melanogaster or homo_sapiens
--rsync-link=<rsync://path/to/dir> Specify an ftp repository to build the database from.
it must be one of the organisms available in ENSEMBLs ftp repository.
And in the same format as its ENSEMBL rsync directoy path.
E.g.:
rsync://ftp.ensembl.org/ensembl/pub/release-81/
rsync://ftp.ensemblgenomes.org/all/pub/protists/current/
rsync://ftp.ensemblgenomes.org/all/pub/plants/current/
rsync://ftp.ensemblgenomes.org/all/pub/fungi/current/
rsync://ftp.ensemblgenomes.org/all/pub/metazoa/current/
target_ident to identify target sequences.
--output-dir=<path/to/dir> - a working directory as unix path to directory.
--parameter-file=<path/to/dir> - a parameter file in cld format as path to file.
--gene-list=<path/to/dir> - a gene list file with ENSEMBL IDs new-line seprated as path to file.
--scoring-module=<path/to/dir> - the path and filename of a file defining a perl scoring function
library_assembly to format a library from an identification folder.
--output-dir=<path/to/dir> - a working directory as unix path to directory.
--parameter-file=<path/to/dir> - a parameter file in cld format as path to file.
--gene-list=<path/to/dir> - a gene list file with ENSEMBL IDs new-line seprated as path to file.
--cov=<int> - Specify the minimum gene coverage as <int> default(15)
--lib-size=<int> - Specify the maximum library size as <int> default(2000)
--library_name=<string> - Prefix for the final library as <string> default(test_lib).
--5-prime=<string> - Define the adapter to be put in 5' before the target site.
default(CTGAGCTCATAGAAGACCTCACC)
--3-prime=<string> - Define the adapter to be put in 3' behind the target site.
default(GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTGGGTCTTCGTTCG)
--cor-5-prime=<string> - Specify if the first 5' baspair should be corrected to a G
may be 1 or 0 default :1.
--input-folder=<path/to/dir> - Specify the input folder for library assembly.
this folder must be prepared by --task= target_ident
--spread-over-transcripts=<string> - should the designs be equally spread oer the different transcripts of the gene
-an be : 1 or 0 (default:0)
end_to_end to perform and end-to-end analysis from target identification to library formatting
--output-dir=<path/to/dir> - a working directory as unix path to directory.
--parameter-file=<path/to/dir> - a parameter file in cld format as path to file.
--gene-list=<path/to/dir> - a gene list file with ENSEMBL IDs new-line seprated as path to file.
--cov=<int> - Specify the minimum gene coverage as <int> default(15)
--lib-size=<int> - Specify the maximum library size as <int> default(2000)
--library_name=<string> - Prefix for the final library as <string> default(test_lib).
--5-prime=<string> - Define the adapter to be put in 5' before the target site.
default(CTGAGCTCATAGAAGACCTCACC)
--3-prime=<string> - Define the adapter to be put in 3' behind the target site.
default(GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTGGGTCTTCGTTCG)
--cor-5-prime=<string> - Specify if the first 5' baspair should be corrected to a G.
--spread-over-transcripts=<string> - should the designs be equally spread oer the different transcripts of the gene
-can be : 1 or 0 (default:1)
--scoring-module=<path/to/dir> - the path and filename of a file defining a perl scoring function
--version - Show version.
--help - Show this message.
};
print (($something{"version"} ? $something{"version_string"} : ''), ($something{"help"} ? $something{"help_string"} : ''));
###################################################################################################################################################################################################
# read in command line options
###################################################################################################################################################################################################
#if(!can_run('bowtie') ){
# print "Cannot find bowtie. Please download and install from\n http://bowtie-bio.sourceforge.net/index.shtml\n";
# print $something{"version_string"}."\n";
# exit;
# }elsif(!can_run('bowtie2')){
# print "Cannot find bowtie2. Please download and install from\n http://bowtie-bio.sourceforge.net/bowtie2/index.shtml\n";
# print $something{"version_string"}."\n";
# exit;
#}
if ($something{"GUI"}) {
###################################################################################################################################################################################################
# define the default parameters for the program
###################################################################################################################################################################################################
#hardcode the choosable Kit presets
%parameter_presets=();
%{$parameter_presets{"strict"}} =(
"preceding"=>"G",
"PAM"=>"NGG",
"min_length"=>20,
"max_length"=>20,
"min_G"=>1,
"max_G"=>80,
"min_A"=>1,
"max_A"=>80,
"min_C"=>1,
"max_C"=>80,
"min_T"=>1,
"max_T"=>80,
"bowtie_version"=>"bowtie",
"offtargetdb"=>"genomeDNA",
"off-targets-allowed"=>5,
"unspecific_leading_bases"=>6,
"edit_distance_allowed"=>2,
"bowtie_mode"=>"very-sensitive",
"ignore_intergenic"=>0,
"sec_off_target"=>0,
"purpose"=>"knockout",
"gene_exclusive"=>1,
"exon_exclusive"=>1,
"CDS_only"=>1,
"specific_exon"=>"any",
"specific_transcript"=>"any",
"exclude_overlapping_genes"=>1,
"CpG_exclusive"=>1,
"ignore_missing_id"=>1,
"kind"=>"single",
"match_info"=>0,
"draw_html_report"=>0,
"max_per_exon"=>40,
"purpose_exclusive"=>1,
"downstream_window"=>50,
"upstream_window"=>50,
"number_of_CDS"=>1,
"minspacerlength"=>15,
"maxspacerlength"=>17,
"crispra_upstream"=>400,
"crispra_downstream"=>50,
"crispri_upstream"=>400,
"crispri_downstream"=>50
);
%{$parameter_presets{"medium"}} =(
"preceding"=>"N",
"PAM"=>"NGG",
"min_length"=>20,
"max_length"=>20,
"min_G"=>1,
"max_G"=>80,
"min_A"=>1,
"max_A"=>80,
"min_C"=>1,
"max_C"=>80,
"min_T"=>1,
"max_T"=>80,
"bowtie_version"=>"bowtie",
"offtargetdb"=>"genomeDNA",
"off-targets-allowed"=>10,
"unspecific_leading_bases"=>1,
"edit_distance_allowed"=>1,
"bowtie_mode"=>"sensitive",
"ignore_intergenic"=>0,
"purpose"=>"knockout",
"gene_exclusive"=>1,
"exon_exclusive"=>1,
"CDS_only"=>0,
"exclude_overlapping_genes"=>0,
"CpG_exclusive"=>1,
"ignore_missing_id"=>1,
"kind"=>"single",
"match_info"=>0,
"draw_html_report"=>0,
"max_per_exon"=>40,
"purpose_exclusive"=>0,
"downstream_window"=>50,
"upstream_window"=>50,
"number_of_CDS"=>3,
"minspacerlength"=>13,
"maxspacerlength"=>19,
"crispra_upstream"=>500,
"crispra_downstream"=>70,
"crispri_upstream"=>500,
"crispri_downstream"=>70
);
%{$parameter_presets{"relaxed"}} =(
"preceding"=>"N",
"PAM"=>"NRG",
"min_length"=>20,
"max_length"=>20,
"min_G"=>1,
"max_G"=>90,
"min_A"=>1,
"max_A"=>90,
"min_C"=>1,
"max_C"=>90,
"min_T"=>1,
"max_T"=>90,
"bowtie_version"=>"bowtie2",
"offtargetdb"=>"gDNA",
"off-targets-allowed"=>20,
"unspecific_leading_bases"=>0,
"edit_distance_allowed"=>0,
"bowtie_mode"=>"fast",
"ignore_intergenic"=>1,
"purpose"=>"knockout",
"gene_exclusive"=>0,
"exon_exclusive"=>0,
"CDS_only"=>0,
"exclude_overlapping_genes"=>0,
"CpG_exclusive"=>0,
"ignore_missing_id"=>0,
"kind"=>"single",
"match_info"=>0,
"draw_html_report"=>0,
"max_per_exon"=>400,
"purpose_exclusive"=>0,
"downstream_window"=>500,
"upstream_window"=>500,
"number_of_CDS"=>10,
"minspacerlength"=>11,
"maxspacerlength"=>21,
"crispra_upstream"=>700,
"crispra_downstream"=>200,
"crispri_upstream"=>700,
"crispri_downstream"=>200);
%{$parameter_presets{"CRISPRi"}} =(
"preceding"=>"G",
"PAM"=>"NRG",
"min_length"=>20,
"max_length"=>20,
"min_G"=>1,
"max_G"=>90,
"min_A"=>1,
"max_A"=>90,
"min_C"=>1,
"max_C"=>90,
"min_T"=>1,
"max_T"=>90,
"bowtie_version"=>"bowtie",
"offtargetdb"=>"gDNA",
"off-targets-allowed"=>5,
"unspecific_leading_bases"=>3,
"edit_distance_allowed"=>2,
"bowtie_mode"=>"very-sensitive",
"ignore_intergenic"=>0,
"sec_off_target"=>0,
"purpose"=>"CRISPRi",
"gene_exclusive"=>0,
"exon_exclusive"=>0,
"CDS_only"=>0,
"specific_exon"=>"any",
"specific_transcript"=>"any",
"exclude_overlapping_genes"=>1,
"CpG_exclusive"=>0,
"ignore_missing_id"=>1,
"kind"=>"single",
"match_info"=>0,
"draw_html_report"=>0,
"max_per_exon"=>40,
"purpose_exclusive"=>1,
"downstream_window"=>50,
"upstream_window"=>50,
"number_of_CDS"=>1,
"minspacerlength"=>15,
"maxspacerlength"=>17,
"crispra_upstream"=>400,
"crispra_downstream"=>50,
"crispri_upstream"=>400,
"crispri_downstream"=>50
);
%{$parameter_presets{"CRISPRa"}} =(
"preceding"=>"G",
"PAM"=>"NRG",
"min_length"=>20,
"max_length"=>20,
"min_G"=>1,
"max_G"=>90,
"min_A"=>1,
"max_A"=>90,
"min_C"=>1,
"max_C"=>90,
"min_T"=>1,
"max_T"=>90,
"bowtie_version"=>"bowtie",
"offtargetdb"=>"gDNA",
"off-targets-allowed"=>5,
"unspecific_leading_bases"=>3,
"edit_distance_allowed"=>2,
"bowtie_mode"=>"very-sensitive",
"ignore_intergenic"=>0,
"sec_off_target"=>0,
"purpose"=>"CRISPRa",
"gene_exclusive"=>0,
"exon_exclusive"=>0,
"CDS_only"=>0,
"specific_exon"=>"any",
"specific_transcript"=>"any",
"exclude_overlapping_genes"=>1,
"CpG_exclusive"=>0,
"ignore_missing_id"=>1,
"kind"=>"single",
"match_info"=>0,
"draw_html_report"=>0,
"max_per_exon"=>40,
"purpose_exclusive"=>1,
"downstream_window"=>50,
"upstream_window"=>50,
"number_of_CDS"=>1,
"minspacerlength"=>15,
"maxspacerlength"=>17,
"crispra_upstream"=>400,
"crispra_downstream"=>50,
"crispri_upstream"=>400,
"crispri_downstream"=>50
);
%{$parameter_presets{"non-coding"}} =(
"preceding"=>"G",
"PAM"=>"NRG",
"min_length"=>20,
"max_length"=>20,
"min_G"=>1,
"max_G"=>90,
"min_A"=>1,
"max_A"=>90,
"min_C"=>1,
"max_C"=>90,
"min_T"=>1,
"max_T"=>90,
"bowtie_version"=>"bowtie",
"offtargetdb"=>"gDNA",
"off-targets-allowed"=>5,
"unspecific_leading_bases"=>3,
"edit_distance_allowed"=>2,
"bowtie_mode"=>"very-sensitive",
"ignore_intergenic"=>0,
"sec_off_target"=>0,
"purpose"=>"non-coding",
"gene_exclusive"=>1,
"exon_exclusive"=>0,
"CDS_only"=>0,
"specific_exon"=>"any",
"specific_transcript"=>"any",
"exclude_overlapping_genes"=>1,
"CpG_exclusive"=>0,
"ignore_missing_id"=>1,
"kind"=>"single",
"match_info"=>0,
"draw_html_report"=>0,
"max_per_exon"=>400,
"purpose_exclusive"=>1,
"downstream_window"=>50,
"upstream_window"=>50,
"number_of_CDS"=>1,
"minspacerlength"=>15,
"maxspacerlength"=>17,
"crispra_upstream"=>400,
"crispra_downstream"=>50,
"crispri_upstream"=>400,
"crispri_downstream"=>50
);
#hardcode the parameter defaults for the following scripts
#general sgRNA properties ;#
#$something{"preceding"}="any" ;#
#$something{"PAM"}="NGG" ;#
$something{"min_length"}=20 ;#
$something{"max_length"}=20 ;#
$something{"min_G"}=1 ;#
$something{"max_G"}=90 ;#
$something{"min_A"}=1 ;#
$something{"max_A"}=90 ;#
$something{"min_C"}=1 ;#
$something{"max_C"}=90 ;#
$something{"min_T"}=1 ;#
$something{"max_T"}=90 ;#
#off-target specific options ;#
#$something{"bowtie_version"}="bowtie" ;#
#$something{"offtargetdb"}="gDNA" ;#
$something{"off-targets-allowed"}=5 ;#
$something{"unspecific_leading_bases"}=5 ;#
$something{"edit_distance_allowed"}=2 ;#
#$something{"bowtie_mode"}="very-sensitive" ;#
$something{"ignore_intergenic"}=0 ;#
$something{"sec_off_target"}=0 ;#
#locus annotation specific options ;#
#$something{"purpose"}="knockout" ;#
$something{"gene_exclusive"}=1 ;#
$something{"exon_exclusive"}=1 ;#
$something{"CDS_only"}=1 ;#
$something{"specific_exon"}="any" ;#
$something{"specific_transcript"}="any" ;#
$something{"exclude_overlapping_genes"}=0 ;#
$something{"CpG_exclusive"}=0 ;#
#input/output options ;#
$something{"databasepath"}="/Users/heigwer/Desktop" ;#
$something{"ref_organism"}="homo_sapiens" ;#
# $something{"data_type"}="ensemble_acc" ;#
$something{"ignore_missing_id"}=1 ;#
# $something{"kind"}="single" ;#
$something{"match_info"}=0 ;#
$something{"draw_html_report"}=0 ;#
$something{"max_per_exon"}=4000 ;#
$something{"out_gff"}=1 ;#
#specific options ;#
$something{"purpose_exclusive"}=0 ;#determines if purpose specific options takes effect
#options for tagging ;#
$something{"retrieve_recomb_matrix"}=0 ;#
$something{"right_homology"}=500 ;#
$something{"left_homology"}=500 ;#
#option for tagging and KO ;#
$something{"downstream_window"}=50 ;#
$something{"upstream_window"}=50 ;#
$something{"number_of_CDS"}=1 ;#
#options for double design after Ran et al. 2014 ;#
$something{"minspacerlength"}=15 ;#
$something{"maxspacerlength"}=17 ;#
#options for CRISPRa/i ;#
$something{"crispra_upstream"}=400 ;#
$something{"crispra_downstream"}=50 ;#
$something{"crispri_upstream"}=400 ;#
$something{"crispri_downstream"}=50 ;#
$something{"coverage"}=15 ;#
$something{"total_lib_size"}=2000 ;#
$something{"library_name"}="test_lib" ;#
$something{"5_adapt"}="CTGAGCTCATAGAAGACCTCACC" ;#
$something{"3_adapt"}="TTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTGGGTCTTCGTTCG";#
$something{"correct_5_prime_G"}=0 ;#
$something{"cover_many_transcripts"}=0 ;#
$something{"sort_by_rank"}=0;
$something{"working_path"}=".";
$something{"custom_score"}=$something{"default_custom_score"}='
##########################################################################################
# Adapted to perl after Doench et al. 2014, NAt. Biotechnology
##########################################################################################
sub {
my $score;
if (length($_[0])==30) {
my %sing_nuc_hash = ("G2"=>-0.275377128,"A3"=>-0.323887456,"C3"=>0.172128871,"C4"=>-0.100666209,"C5"=>-0.20180294,
"G5"=>0.245956633,"A6"=>0.036440041,"C6"=>0.098376835,"C7"=>-0.741181291,
"G7"=>-0.393264397,"A12"=>-0.466099015,"A15"=>0.085376945,"C15"=>-0.013813972,
"A16"=>0.272620512,"C16"=>-0.119022648,"T16"=>-0.285944222,"A17"=>0.097454592,
"G17"=>-0.17554617,"C18"=>-0.345795451,"G18"=>-0.678096426,"A19"=>0.22508903,
"C19"=>-0.507794051,"G20"=>-0.417373597,"T20"=>-0.054306959,"G21"=>0.379899366,
"T21"=>-0.090712644,"C22"=>0.057823319,"T22"=>-0.530567296,"T23"=>-0.877007428,
"C24"=>-0.876235846,"G24"=>0.278916259,"T24"=>-0.403102218,"A25"=>-0.077300704,
"C25"=>0.287935617,"T25"=>-0.221637217,"G28"=>-0.689016682,"T28"=>0.117877577,
"C29"=>-0.160445304,"G30"=>0.386342585);
my %dinuc_hash = ("GT2"=>-0.625778696,"GC5"=>0.300043317,"AA6"=>-0.834836245,"TA6"=>0.760627772,"GG7"=>-0.490816749,
"GG12"=>-1.516907439,"TA12"=>0.7092612,"TC12"=>0.496298609,"TT12"=>-0.586873894,"GG13"=>-0.334563735,
"GA14"=>0.76384993,"GC14"=>-0.53702517,"TG17"=>-0.798146133,"GG19"=>-0.66680873,"TC19"=>0.353183252,
"CC20"=>0.748072092,"TG20"=>-0.367266772,"AC21"=>0.568209132,"CG21"=>0.329072074,"GA21"=>-0.836456755,
"GG21"=>-0.782207584,"TC22"=>-1.029692957,"CG23"=>0.856197823,"CT23"=>-0.463207679,"AA24"=>-0.579492389,
"AG24"=>0.649075537,"AG25"=>-0.077300704,"CG25"=>0.287935617,"TG25"=>-0.221637217,"GT27"=>0.117877577,
"GG29"=>-0.697740024);
my $gc = ( substr($_[0],4,20) =~ tr/GC/GC/);
if ($gc < 10){
$score=0.597636154+(abs($gc-10)*-0.202625894)
}else{
$score=0.597636154+(($gc-10)*-0.166587752)
}
foreach my $i (0..29){
my $key = substr($_[0],$i,1).($i+1);
if ($sing_nuc_hash{$key}) {
$score+=$sing_nuc_hash{$key};
}
if($i<29){
$key =substr($_[0],$i,2).($i+1);
if ($dinuc_hash{$key}){
$score+=$dinuc_hash{$key};
}
}
}
return(1/(1+exp(-$score)))
#code
}else{
return(0);
}
};';
###################################################################################################################################################################################################
###################################################################################################################################################################################################
open( $temp_file, ">", "temp") or die $!;
print $temp_file ';=================================================
; Theme : snow
; Author : Djibril Ousmanou
; Date : 01/01/2011 00:00:00
;=================================================
[Balloon]
-background: #FFFFFF
-foreground: #0047B9
[BrowseEntry]
-background: #FFFFFF
-disabledbackground: #FFFFFF
-disabledforeground: #5F5E5E
-foreground: #0047B9
[Button]
-activebackground: #A8A8A7
-activeforeground: #0047B9
-background: #A8A8A7
-disabledforeground: #46562C
-foreground: #0047B9
[Canvas]
-background: #FFFFFF
[Checkbutton]
-activebackground: #FFFFFF
-activeforeground: #0047B9
-background: #FFFFFF
-disabledforeground: #0047B9
-foreground: #0047B9
-highlightbackground: #FFFFFF
-selectcolor: #0047B9
[ColoredButton]
-autofit: 1
-background: #FFFFFF
-highlightbackground: #FFFFFF
[DirTree]
-background: #FFFFFF
-foreground: #0047B9
-highlightbackground: #FFFFFF
-highlightcolor: #FFFFFF
-selectbackground: #A8A8A7
-selectforeground: #0047B9
[Entry]
-background: #FFFFFF
-disabledbackground: #FFFFFF
-disabledforeground: #5F5E5E
-foreground: #0047B9
-highlightbackground: #FFFFFF
-insertbackground: #0047B9
-readonlybackground: #FFFFFF
-selectbackground: #A8A8A7
[EntryCheck]
-background: #FFFFFF
-disabledbackground: #FFFFFF
-disabledforeground: #5F5E5E
-foreground: #0047B9
-highlightbackground: #FFFFFF
-insertbackground: #0047B9
-readonlybackground: #FFFFFF
-selectbackground: #A8A8A7
[Frame]
-background: #FFFFFF
-highlightbackground: #FFFFFF
[HList]
-background: #FFFFFF
-foreground: #0047B9
-highlightbackground: #FFFFFF
-highlightcolor: #FFFFFF
-selectbackground: #A8A8A7
-selectforeground: #0047B9
[LabEntry]
-background: #FFFFFF
-disabledbackground: #FFFFFF
-disabledforeground: #5F5E5E
[LabFrame]
-background: #FFFFFF
-foreground: #0047B9
[Label]
-background: #FFFFFF
-foreground: #0047B9
-highlightbackground: #FFFFFF
[Labelframe]
-background: #FFFFFF
-foreground: #0047B9
-highlightbackground: #FFFFFF
-highlightcolor: #FFFFFF
[Listbox]
-background: #FFFFFF
-foreground: #0047B9
-highlightbackground: #FFFFFF
-highlightcolor: #FFFFFF
-selectbackground: #A8A8A7
-selectforeground: #0047B9
[MainWindow]
-background: #FFFFFF
-highlightbackground: #FFFFFF
-highlightcolor: #FFFFFF
[Menu]
-activebackground: #FFFFFF
-activeforeground: #0047B9
-background: #FFFFFF
-foreground: #0047B9
-selectcolor: #FFFFFF
[Menubutton]
-activebackground: #A8A8A7
-activeforeground: #0047B9
-background: #A8A8A7
-disabledforeground: #46562C
-foreground: #0047B9
[NoteBook]
-background: #FFFFFF
-backpagecolor: #FFFFFF
-disabledforeground: #5F5E5E
-focuscolor: #FFFFFF
-foreground: #0047B9
-inactivebackground: #FFFFFF
[Optionmenu]
-activebackground: #A8A8A7
-activeforeground: #0047B9
-background: #A8A8A7
-borderwidth: 0
-disabledforeground: #46562C
-foreground: #0047B9
[ProgressBar]
-colors: 0
-colors: #00FF7B
-troughcolor: #FFFFFF
[ProgressBarPlus]
-colors: 0
-colors: #00FF7B
-troughcolor: #FFFFFF
[ROText]
-background: #FFFFFF
-foreground: #0047B9
-insertbackground: #0047B9
[Radiobutton]
-activebackground: #FFFFFF
-activeforeground: #0047B9
-background: #FFFFFF
-disabledforeground: #0047B9
-foreground: #0047B9
-highlightbackground: #FFFFFF
-selectcolor: #FFFFFF
[Scale]
-activebackground: #A8A8A7
-background: #FFFFFF
-foreground: #0047B9
-highlightbackground: #FFFFFF
-highlightcolor: #FFFFFF
-troughcolor: #FFFFFF
[Spinbox]
-background: #FFFFFF
-buttonbackground: #A8A8A7
-foreground: #0047B9
-readonlybackground: #FFFFFF
[TList]
-background: #FFFFFF
-foreground: #0047B9
-highlightcolor: #FFFFFF
-selectbackground: #A8A8A7
-selectforeground: #0047B9
[Table]
-background: #FFFFFF
-highlightbackground: #FFFFFF
[Text]
-background: #FFFFFF
-foreground: #0047B9
-insertbackground: #0047B9
[TextUndo]
-background: #FFFFFF
-foreground: #0047B9
-insertbackground: #0047B9
[Toplevel]
-background: #FFFFFF
-highlightbackground: #FFFFFF
-highlightcolor: #FFFFFF
[Tree]
-background: #FFFFFF
-foreground: #0047B9
-highlightbackground: #FFFFFF
-highlightcolor: #FFFFFF
-selectbackground: #A8A8A7
-selectforeground: #0047B9
';
close($temp_file);
# Set it to e frame widget
###################################################################################################################################################################################################
# create a graphical user interface (GUI) to render inputs and output in a human radble form
###################################################################################################################################################################################################
$mw = MainWindow->new( -title => "cld parameter", );
$mw->minsize( 500, 500 );
$higher_order_functions=$mw->NoteBook();
$make_database = $higher_order_functions ->add('page1', -label => 'Genome Data')->NoteBook();
$gene_list = $higher_order_functions ->add('page2', -label => 'Gene List')->NoteBook();
$end_to_end = $higher_order_functions ->add('page3', -label => 'Design Parameter')->NoteBook();
$higher_order_functions -> grid(-row=>1,-column=>1,-sticky=>"nw");
$gene_list -> grid(-row=>1,-column=>1,-columnspan=>2,-sticky=>"nw");
$end_to_end -> grid(-row=>1,-column=>1,-sticky=>"nw");
$make_database -> grid(-row=>1,-column=>1,-sticky=>"nw");
$lab_databasepath = $make_database -> Button(-text=>"Select folder for \ndeposition genome data",
-command=>\&choosepathtoindex,
-width=>20,-anchor =>"w"); #make the button to trigger the file choosing function
$databasepath_entry = $make_database -> PathEntry(
-textvariable=>\$something{"databasepath"},
-width=>40,
-background=>"white"
); #make the Text/Path entry widget for assisted entry
sub choosepathtoindex{ #function triggered by the button to choose an openable existing file
$something{"databasepath"}=$mw->chooseDirectory(-title=>"Please choose an folder containing organism data",-initialdir=>"~"); #open the file opening dialog
$something{"ref_organism"}=$ref_organism_entry->get('1.0','end-1c');
if ($something{"databasepath"}=~/[\s]+/) {
$something{"databasepath"}="";
$databasepath_entry -> configure( -background => "#d73027" );
$make_database->messageBox(
-icon => 'error',
-type => 'ok',
-title => 'Error',
-message => 'Folder names may not contain characters other than A-Za-z0-9_-:',
);
}
if(-d $something{"databasepath"}."/".$something{"ref_organism"}){
$databasepath_entry -> configure( -background => "#FFFFFF" );
$ref_organism_entry -> configure( -background => "#FFFFFF" );
}else{
$databasepath_entry -> configure( -background => "#d73027" );
$ref_organism_entry -> configure( -background => "#d73027" );
}
}
$lab_ref_organism = $make_database -> Label(-text=>"please type the name of the \nreference organism as given in the database\n(e.g. drosophila_melanogaster)",-anchor =>"w"); #create a label object
$ref_organism_entry = $make_database -> Text( -height=>1,
-width=>40); #create a text entry object
$ref_organism_entry->bind('<<Modified>>'=>sub {
if($ref_organism_entry->editModified) {
$something{"ref_organism"}=$something{"organism_db"}=$ref_organism_entry->get('1.0','end-1c');
if(-d $something{"databasepath"}."/".$something{"ref_organism"}){
$databasepath_entry -> configure( -background => "#FFFFFF" );
$ref_organism_entry -> configure( -background => "#FFFFFF" );
}else{
$databasepath_entry -> configure( -background => "#d73027" );
$ref_organism_entry -> configure( -background => "#d73027" );
}
$ref_organism_entry->editModified(0)
}
});
#$ref_organism_entry->insert('1.0',$something{"ref_organism"});
$opt_rsync=$make_database->Optionmenu(
-options => [
["ensembl-pub"=>"rsync://ftp.ensembl.org/ensembl/pub/release-81/"],
["ensemblgenomes-protists"=>"rsync://ftp.ensemblgenomes.org/all/pub/protists/current/"],
["ensemblgenomes-plants"=>"rsync://ftp.ensemblgenomes.org/all/pub/plants/current/"],
["ensemblgenomes-fungi"=>"rsync://ftp.ensemblgenomes.org/all/pub/fungi/current/"],
["ensemblgenomes-metazoa"=>"rsync://ftp.ensemblgenomes.org/all/pub/metazoa/current/"]
],
-variable => \$something{"rsync_link"}
);
$opt_precalc=$make_database->Optionmenu(
-options => [
["drosophila_melanogaster.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/drosophila_melanogaster.tar.gz"],
["aedes_aegypti.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/aedes_aegypti.tar.gz"],
["anopheles_darlingi.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/anopheles_darlingi.tar.gz"],
["anopheles_gambiae.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/anopheles_gambiae.tar.gz"],
["arabidopsis_thaliana.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/arabidopsis_thaliana.tar.gz"],
["aspergillus_nidulans.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/aspergillus_nidulans.tar.gz"],
["aspergillus_niger.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/aspergillus_niger.tar.gz"],
["brachypodium_distachyon.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/brachypodium_distachyon.tar.gz"],
["chlamydomonas_reinhardtii.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/chlamydomonas_reinhardtii.tar.gz"],
["ciona_intestinalis.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/ciona_intestinalis.tar.gz"],
["cricetulus_griseus.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/cricetulus_griseus.tar.gz"],
["dictyostelium_discoideum.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/dictyostelium_discoideum.tar.gz"],
["drosophila_pseudoobscura.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/drosophila_pseudoobscura.tar.gz"],
["emiliania_huxleyi.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/emiliania_huxleyi.tar.gz"],
["homo_sapiens.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/homo_sapiens.tar.gz"],
["hordeum_vulgare.tar.gz"=>"http://www.dkfz.de/signaling/crispr-downloads/DATABASES/hordeum_vulgare.tar.gz"],