From cc2325031a60709be32b05f512209e3cd5e1f7ba Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ren=C3=A9=20Wiegandt?= Date: Tue, 18 Dec 2018 08:00:39 -0500 Subject: [PATCH 01/33] Bugfix in bed_to_fasta.R: Get last and second last instead of fixed indices --- bin/bed_to_fasta.R | 62 +++++++++++++++++++++++++--------------------- 1 file changed, 34 insertions(+), 28 deletions(-) diff --git a/bin/bed_to_fasta.R b/bin/bed_to_fasta.R index 84910f3..8af8516 100644 --- a/bin/bed_to_fasta.R +++ b/bin/bed_to_fasta.R @@ -1,28 +1,34 @@ -#!/usr/bin/env Rscript - -# Splitting BED-files depending on their cluster. -# The Sequences of each cluster are writen as an FASTA-file. -# @parameter bedInput BED-file with sequences and cluster-id as columns: Sequence: Column 7; ID:Column 8 -# @parameter prefix prefix for filenames -# @parameter min_seq min. number of sequences per cluster - -args = commandArgs(trailingOnly = TRUE) - -bedInput <- args[1] -prefix <- args[2] -min_seq <- args[3] - -bed <- data.table::fread(bedInput, header = FALSE, sep = "\t") - -clusters <- split(bed, bed$V11, sorted = TRUE, flatten = FALSE) # <---- Cluster column -discard <- lapply(1:length(clusters), function(i){ - clust <- as.data.frame(clusters[i]) - print(nrow(clust)) - if (nrow(clust) >= as.numeric(min_seq) ) { - sequences <- as.list(clust[[10]]) # <---- sequenze column - outfile <- paste0(prefix,"_cluster_",i,".FASTA") - seqinr::write.fasta(sequences = sequences, names = clust[[4]], file.out = outfile, as.string = TRUE) # <---- Name column - } else { - print(paste0("Cluster: ",i," is to small")) - } -}) +#!/usr/bin/env Rscript + +#' Splitting BED-files depending on their cluster. +#' The Sequences of each cluster are writen as an FASTA-file. +#' @parameter bedInput BED-file with sequences and cluster-id as columns: Sequence: Column 7; ID:Column 8 +#' @parameter prefix prefix for filenames +#' @parameter min_seq min. number of sequences per cluster +#' +#' @author René Wiegandt +#' @contact rene.wiegandt(at)mpi-bn.mpg.de + +args = commandArgs(trailingOnly = TRUE) + +bedInput <- args[1] +prefix <- args[2] +min_seq <- args[3] + +bed <- data.table::fread(bedInput, header = FALSE, sep = "\t") + +# Get last column of data.table, which refers to the cluster, as a vector. +cluster_no <- as.vector(bed[[ncol(bed)]]) + +clusters <- split(bed, cluster_no, sorted = TRUE, flatten = FALSE) # <---- Cluster column + +discard <- lapply(1:length(clusters), function(i){ + clust <- as.data.frame(clusters[i]) + if (nrow(clust) >= as.numeric(min_seq) ) { + sequences <- as.list(clust[[ncol(clust) - 1]]) # <---- sequenze column + outfile <- paste0(prefix,"_cluster_",i - 1,".FASTA") + seqinr::write.fasta(sequences = sequences, names = clust[[4]], file.out = outfile, as.string = TRUE) # <---- Name column + } else { + print(paste0("Cluster: ",i," is to small")) + } +}) From 62f6f3f1e95ad6038567619a3f846203f1e92751 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ren=C3=A9=20Wiegandt?= Date: Tue, 18 Dec 2018 08:10:13 -0500 Subject: [PATCH 02/33] bed_to_fasta.R: Improved documentation --- bin/bed_to_fasta.R | 19 +++++++++++++------ 1 file changed, 13 insertions(+), 6 deletions(-) diff --git a/bin/bed_to_fasta.R b/bin/bed_to_fasta.R index 8af8516..dce3839 100644 --- a/bin/bed_to_fasta.R +++ b/bin/bed_to_fasta.R @@ -2,9 +2,10 @@ #' Splitting BED-files depending on their cluster. #' The Sequences of each cluster are writen as an FASTA-file. -#' @parameter bedInput BED-file with sequences and cluster-id as columns: Sequence: Column 7; ID:Column 8 -#' @parameter prefix prefix for filenames -#' @parameter min_seq min. number of sequences per cluster +#' @param bedInput BED-file with sequences and cluster-id as last two columns: +#' Sequence: second last column; Cluster ID: last column +#' @param prefix prefix for filenames +#' @param min_seq min. number of sequences per cluster #' #' @author René Wiegandt #' @contact rene.wiegandt(at)mpi-bn.mpg.de @@ -20,14 +21,20 @@ bed <- data.table::fread(bedInput, header = FALSE, sep = "\t") # Get last column of data.table, which refers to the cluster, as a vector. cluster_no <- as.vector(bed[[ncol(bed)]]) -clusters <- split(bed, cluster_no, sorted = TRUE, flatten = FALSE) # <---- Cluster column +# Split data.table bed on its last column (cluster_no) into list of data.frames +clusters <- split(bed, cluster_no, sorted = TRUE, flatten = FALSE) +# For each data.frame(cluster) in list clusters: discard <- lapply(1:length(clusters), function(i){ clust <- as.data.frame(clusters[i]) + # Filter data.tables(clusters), which are to small if (nrow(clust) >= as.numeric(min_seq) ) { - sequences <- as.list(clust[[ncol(clust) - 1]]) # <---- sequenze column + # Get second last column, which contains the nucleotide sequences + sequences <- as.list(clust[[ncol(clust) - 1]]) + # Create filename outfile <- paste0(prefix,"_cluster_",i - 1,".FASTA") - seqinr::write.fasta(sequences = sequences, names = clust[[4]], file.out = outfile, as.string = TRUE) # <---- Name column + # Write fasta file + seqinr::write.fasta(sequences = sequences, names = clust[[4]], file.out = outfile, as.string = TRUE) } else { print(paste0("Cluster: ",i," is to small")) } From cf9dcd84983c4825d52b38c28cc7314c67b367ae Mon Sep 17 00:00:00 2001 From: renewiegandt Date: Wed, 19 Dec 2018 12:16:13 +0100 Subject: [PATCH 03/33] bed_to_fasta.R: Imporved parametercalling with optparse --- bin/bed_to_fasta.R | 99 +++++++++++++++++++++++++++------------------- 1 file changed, 58 insertions(+), 41 deletions(-) diff --git a/bin/bed_to_fasta.R b/bin/bed_to_fasta.R index dce3839..56ac9dc 100644 --- a/bin/bed_to_fasta.R +++ b/bin/bed_to_fasta.R @@ -1,41 +1,58 @@ -#!/usr/bin/env Rscript - -#' Splitting BED-files depending on their cluster. -#' The Sequences of each cluster are writen as an FASTA-file. -#' @param bedInput BED-file with sequences and cluster-id as last two columns: -#' Sequence: second last column; Cluster ID: last column -#' @param prefix prefix for filenames -#' @param min_seq min. number of sequences per cluster -#' -#' @author René Wiegandt -#' @contact rene.wiegandt(at)mpi-bn.mpg.de - -args = commandArgs(trailingOnly = TRUE) - -bedInput <- args[1] -prefix <- args[2] -min_seq <- args[3] - -bed <- data.table::fread(bedInput, header = FALSE, sep = "\t") - -# Get last column of data.table, which refers to the cluster, as a vector. -cluster_no <- as.vector(bed[[ncol(bed)]]) - -# Split data.table bed on its last column (cluster_no) into list of data.frames -clusters <- split(bed, cluster_no, sorted = TRUE, flatten = FALSE) - -# For each data.frame(cluster) in list clusters: -discard <- lapply(1:length(clusters), function(i){ - clust <- as.data.frame(clusters[i]) - # Filter data.tables(clusters), which are to small - if (nrow(clust) >= as.numeric(min_seq) ) { - # Get second last column, which contains the nucleotide sequences - sequences <- as.list(clust[[ncol(clust) - 1]]) - # Create filename - outfile <- paste0(prefix,"_cluster_",i - 1,".FASTA") - # Write fasta file - seqinr::write.fasta(sequences = sequences, names = clust[[4]], file.out = outfile, as.string = TRUE) - } else { - print(paste0("Cluster: ",i," is to small")) - } -}) +#!/usr/bin/env Rscript +library("optparse") + +option_list <- list( + make_option(opt_str = c("-i", "--input"), default = NULL, help = "Input bed-file. Second last column must be sequences and last column must be the cluster_id.", metavar = "character"), + make_option(opt_str = c("-p", "--prefix"), default = "" , help = "Prefix for file names. Default = '%default'", metavar = "character"), + make_option(opt_str = c("-m", "--min_seq"), default = 100, help = "Minimum amount of sequences in clusters. Default = %default", metavar = "integer") +) + +opt_parser <- OptionParser(option_list = option_list, + description = "Convert BED-file to one FASTA-file per cluster") + +opt <- parse_args(opt_parser) + +#' Splitting BED-files depending on their cluster. +#' The Sequences of each cluster are writen as an FASTA-file. +#' @param bedInput BED-file with sequences and cluster-id as last two columns: +#' Sequence: second last column; Cluster ID: last column +#' @param prefix prefix for filenames +#' @param min_seq min. number of sequences per cluster +#' +#' @author René Wiegandt +#' @contact rene.wiegandt(at)mpi-bn.mpg.de +bed_to_fasta <- function(bedInput, prefix = "", min_seq = 100){ + + if(is.null(bedInput)){ + stop("ERROR: Input parameter cannot be null! Please specify the input parameter.") + } + + bed <- data.table::fread(bedInput, header = FALSE, sep = "\t") + + # Get last column of data.table, which refers to the cluster, as a vector. + cluster_no <- as.vector(bed[[ncol(bed)]]) + + # Split data.table bed on its last column (cluster_no) into list of data.frames + clusters <- split(bed, cluster_no, sorted = TRUE, flatten = FALSE) + + # For each data.frame(cluster) in list clusters: + discard <- lapply(1:length(clusters), function(i){ + clust <- as.data.frame(clusters[i]) + # Filter data.tables(clusters), which are to small + if (nrow(clust) >= as.numeric(min_seq) ) { + # Get second last column, which contains the nucleotide sequences + sequences <- as.list(clust[[ncol(clust) - 1]]) + # Create filename + outfile <- paste0(prefix,"_cluster_",i - 1,".FASTA") + # Write fasta file + seqinr::write.fasta(sequences = sequences, names = clust[[4]], file.out = outfile, as.string = TRUE) + } else { + print(paste0("Cluster: ",i," is to small")) + } + }) +} + +# run function bed_to_fasta with given parameteres if not in interactive context (e.g. run from shell) +if (!interactive()) { + bed_to_fasta(opt$input, opt$prefix, opt$min_seq) +} \ No newline at end of file From 6d5c604c2950e90ab7f12bd4822f8e2e5cafa0f9 Mon Sep 17 00:00:00 2001 From: renewiegandt Date: Wed, 19 Dec 2018 12:18:08 +0100 Subject: [PATCH 04/33] adaption of pipeline.nf to changes in bed_to_fasta.R --- pipeline.nf | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/pipeline.nf b/pipeline.nf index a39616a..463a790 100644 --- a/pipeline.nf +++ b/pipeline.nf @@ -283,7 +283,7 @@ process reduce_bed { """ Rscript ${path_bin}/reduce_bed.R -i ${bed} -k ${params.kmer} -m ${params.aprox_motif_len} -o ${name}_reduced.bed -t ${params.threads} -f ${params.motif_occurence} -s ${params.min_seq_length} """ -} +}String /* @@ -324,7 +324,7 @@ process bed_to_clustered_fasta { script: """ - Rscript ${path_bin}/bed_to_fasta.R ${bed} ${name} ${params.min_seq} + Rscript ${path_bin}/bed_to_fasta.R -i ${bed} -p ${name} -m ${params.min_seq} """ } From ce5287174ca4ddee0e8bc54fca3eb6a18378b8a4 Mon Sep 17 00:00:00 2001 From: renewiegandt Date: Wed, 19 Dec 2018 12:47:00 +0100 Subject: [PATCH 05/33] Refactoring --- bin/bed_to_fasta.R | 2 +- pipeline.nf | 2 +- 2 files changed, 2 insertions(+), 2 deletions(-) diff --git a/bin/bed_to_fasta.R b/bin/bed_to_fasta.R index 56ac9dc..bb4d3dc 100644 --- a/bin/bed_to_fasta.R +++ b/bin/bed_to_fasta.R @@ -55,4 +55,4 @@ bed_to_fasta <- function(bedInput, prefix = "", min_seq = 100){ # run function bed_to_fasta with given parameteres if not in interactive context (e.g. run from shell) if (!interactive()) { bed_to_fasta(opt$input, opt$prefix, opt$min_seq) -} \ No newline at end of file +} diff --git a/pipeline.nf b/pipeline.nf index 463a790..55b7c26 100644 --- a/pipeline.nf +++ b/pipeline.nf @@ -283,7 +283,7 @@ process reduce_bed { """ Rscript ${path_bin}/reduce_bed.R -i ${bed} -k ${params.kmer} -m ${params.aprox_motif_len} -o ${name}_reduced.bed -t ${params.threads} -f ${params.motif_occurence} -s ${params.min_seq_length} """ -}String +} /* From 98985d13512ede1d8c352207ea002d84522a2766 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Wed, 19 Dec 2018 13:47:43 +0100 Subject: [PATCH 06/33] refactoring; renamed reduce_bed to reduce_sequence --- bin/{reduce_bed.R => reduce_sequence.R} | 38 ++++++++++++++----------- 1 file changed, 21 insertions(+), 17 deletions(-) rename bin/{reduce_bed.R => reduce_sequence.R} (84%) diff --git a/bin/reduce_bed.R b/bin/reduce_sequence.R similarity index 84% rename from bin/reduce_bed.R rename to bin/reduce_sequence.R index 2f8e8a2..586b074 100644 --- a/bin/reduce_bed.R +++ b/bin/reduce_sequence.R @@ -9,32 +9,35 @@ option_list <- list( make_option(opt_str = c("-t", "--threads"), default = 1, help = "Number of threads to use. Use 0 for all available cores. Default = %default", metavar = "integer"), make_option(opt_str = c("-c", "--clean"), default = TRUE, help = "Delete all temporary files. Default = %default", metavar = "logical"), make_option(opt_str = c("-s", "--min_seq_length"), default = NULL, help = "Remove sequences below this length. Defaults to the maximum value of motif and kmer and can not be lower.", metavar = "integer", type = "integer"), - make_option(opt_str = c("-n", "--minoverlap_kmer"), default = NULL, help = "Minimum required overlap between kmer to merge kmer. Used to create reduced sequence ranges. Can not be greater than kmer length. Default = kmer - 1", metavar = "integer", type = "integer"), + make_option(opt_str = c("-n", "--minoverlap_kmer"), default = NULL, help = "Minimum required overlap between kmer. Used to create reduced sequence ranges out of merged kmer. Can not be greater than kmer length. Default = kmer - 1", metavar = "integer", type = "integer"), make_option(opt_str = c("-v", "--minoverlap_motif"), default = NULL, help = "Minimum required overlap between motif and kmer to consider kmer significant. Used for kmer cutoff calculation. Can not be greater than motif and kmer length. Default = ceiling(motif / 2)", metavar = "integer", type = "integer"), - make_option(opt_str = c("-f", "--motif_occurence"), default = 1, help = "Number of motifs per sequence any value above 0. Default = %default.", metavar = "double") + make_option(opt_str = c("-f", "--motif_occurence"), default = 1, help = "Define how many motifs are expected per sequence. This value is used during kmer cutoff calculation. Default = %default meaning that there should be approximately one motif per sequence.", metavar = "double") ) opt_parser <- OptionParser(option_list = option_list, - description = "Reduce sequences to frequent regions.") + description = "Reduces each sequence to its most frequent region.") opt <- parse_args(opt_parser) -#' Reduce bed file to conserved regions +#' Reduces each sequence to its most frequent region. #' -#' @param input bed file -#' @param kmer Length of kmer. -#' @param motif Estimated motif length. -#' @param output Output file -#' @param threads Number of threads. Default = 1. 0 for all cores. +#' @param input Input bed-file. Last column must be sequences. +#' @param kmer Kmer length. Default = 10 +#' @param motif Estimated motif length. Default = 10 +#' @param output Output file. Default = reduced.bed +#' @param threads Number of threads to use. Default = 1. Use 0 for all cores. #' @param clean Delete all temporary files. -#' @param minoverlap_kmer Minimum required overlap between kmer to merge kmer. Used to create reduced sequence ranges. Can not be greater than kmer length. Default = kmer - 1 +#' @param minoverlap_kmer Minimum required overlap between kmer. Used to create reduced sequence ranges out of merged kmer. Can not be greater than kmer length . Default = kmer - 1 #' @param minoverlap_motif Minimum required overlap between motif and kmer to consider kmer significant. Used for kmer cutoff calculation. Can not be greater than motif and kmer length. Default = ceiling(motif / 2) -#' @param min_seq_length Must be greater or equal to kmer and motif. Default = max(c(motif, kmer)). +#' @param min_seq_length Remove sequences below this length. Defaults to the maximum value of motif and kmer and can not be lower. #' @param motif_occurence Define how many motifs are expected per sequence. This value is used during kmer cutoff calculation. Default = 1 meaning that there should be approximately one motif per sequence. #' -#' @return reduced bed #' TODO check whether jellyfish is installed -reduce_bed <- function(input, kmer = 10, motif = 10, output = "reduced.bed", threads = NULL, clean = TRUE, minoverlap_kmer = kmer - 1, minoverlap_motif = ceiling(motif / 2), min_seq_length = max(c(motif, kmer)), motif_occurence = 1) { +reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed", threads = NULL, clean = TRUE, minoverlap_kmer = kmer - 1, minoverlap_motif = ceiling(motif / 2), min_seq_length = max(c(motif, kmer)), motif_occurence = 1) { + if (missing(input)) { + stop("No input specified! Please forward a valid bed-file.") + } + # get number of available cores if (threads == 0) { threads <- parallel::detectCores() @@ -117,12 +120,12 @@ reduce_bed <- function(input, kmer = 10, motif = 10, output = "reduced.bed", thr merged[, sequence := stringr::str_sub(sequence, relative_start, relative_end)] # bed files count from 0 - merged[, `:=`(relative_start = relative_start - 1, relative_end = relative_end - 1)] + merged[, data.table::`:=`(relative_start = relative_start - 1, relative_end = relative_end - 1)] # change start end location - merged[, `:=`(start = start + relative_start, end = start + relative_end)] + merged[, data.table::`:=`(start = start + relative_start, end = start + relative_end)] # clean table - merged[, `:=`(relative_start = NULL, relative_end = NULL, width = NULL)] + merged[, data.table::`:=`(relative_start = NULL, relative_end = NULL, width = NULL)] if (clean) { file.remove(fasta_file, count_output_binary, mer_count_table) @@ -171,6 +174,7 @@ significant_kmer <- function(bed, kmer, motif, minoverlap = ceiling(motif / 2), reduce_kmer <- function(kmer, significant) { data.table::setorderv(kmer, cols = names(kmer)[2], order = -1) + # TODO don't use 'V2' kmer[, cumsum := cumsum(V2)] return(kmer[cumsum <= significant]) @@ -255,5 +259,5 @@ if (!interactive()) { pbo <- pbapply::pboptions(type = "timer") # remove last parameter (help param) params <- opt[-length(opt)] - do.call(reduce_bed, args = params) + do.call(reduce_sequence, args = params) } From e0b9d38a9dc5cf978eed2fef436ee7be8ed2b0c9 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Wed, 19 Dec 2018 14:02:54 +0100 Subject: [PATCH 07/33] check whether jellyfish is installed --- bin/reduce_sequence.R | 5 ++++- 1 file changed, 4 insertions(+), 1 deletion(-) diff --git a/bin/reduce_sequence.R b/bin/reduce_sequence.R index 586b074..69fc764 100644 --- a/bin/reduce_sequence.R +++ b/bin/reduce_sequence.R @@ -32,8 +32,11 @@ opt <- parse_args(opt_parser) #' @param min_seq_length Remove sequences below this length. Defaults to the maximum value of motif and kmer and can not be lower. #' @param motif_occurence Define how many motifs are expected per sequence. This value is used during kmer cutoff calculation. Default = 1 meaning that there should be approximately one motif per sequence. #' -#' TODO check whether jellyfish is installed reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed", threads = NULL, clean = TRUE, minoverlap_kmer = kmer - 1, minoverlap_motif = ceiling(motif / 2), min_seq_length = max(c(motif, kmer)), motif_occurence = 1) { + if (system("jellyfish count --version") != 0) { + stop("Required programm jellyfish not found! Please check whether it is installed.") + } + if (missing(input)) { stop("No input specified! Please forward a valid bed-file.") } From 17308687327a8cd17e0b7af3916a58073152f344 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Wed, 19 Dec 2018 14:06:13 +0100 Subject: [PATCH 08/33] reduce_bed renamed to reduce_sequence --- pipeline.nf | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/pipeline.nf b/pipeline.nf index a39616a..db3bbdf 100644 --- a/pipeline.nf +++ b/pipeline.nf @@ -21,7 +21,7 @@ params.max_size_fp=100 //clustering - //reduce_bed + //reduce_sequence params.kmer=10 params.aprox_motif_len=10 params.motif_occurence=1 @@ -268,7 +268,7 @@ process overlap_with_known_TFBS { /* */ -process reduce_bed { +process reduce_sequence { conda "${path_env}" echo true publishDir "${params.out}/cluster/reduced_bed/", mode: 'copy' @@ -281,7 +281,7 @@ process reduce_bed { script: """ - Rscript ${path_bin}/reduce_bed.R -i ${bed} -k ${params.kmer} -m ${params.aprox_motif_len} -o ${name}_reduced.bed -t ${params.threads} -f ${params.motif_occurence} -s ${params.min_seq_length} + Rscript ${path_bin}/reduce_sequence.R -i ${bed} -k ${params.kmer} -m ${params.aprox_motif_len} -o ${name}_reduced.bed -t ${params.threads} -f ${params.motif_occurence} -s ${params.min_seq_length} """ } From 3c4f733eb627dce9c2e8566f617dbe7dbf58a1fd Mon Sep 17 00:00:00 2001 From: renewiegandt Date: Wed, 19 Dec 2018 14:40:59 +0100 Subject: [PATCH 09/33] get_best_motif.py: fixed bug which caused to print motif header as last line + improved documentation --- bin/get_best_motif.py | 54 ++++++++++++++++++++++++++++++++++++------- 1 file changed, 46 insertions(+), 8 deletions(-) diff --git a/bin/get_best_motif.py b/bin/get_best_motif.py index cc24949..37eccc5 100644 --- a/bin/get_best_motif.py +++ b/bin/get_best_motif.py @@ -1,26 +1,64 @@ -# parses arguments using argparse -# @return args list of all parameters +''' +parses arguments using argparse +@return args list of all parameters +''' def parse_arguments(): - parser = argparse.ArgumentParser() - parser.add_argument("meme", help="Path to meme file") + parser = argparse.ArgumentParser(description='A script to convert from GLAM2 output to MEME-format and parsing only the [num] first motifs from file to the output.') + parser.add_argument("meme", help="Path to 'meme' file generated by GLAM2") parser.add_argument("output", help="Output file") parser.add_argument("num", help="Number of motifs parsed from file") args = parser.parse_args() return args -# write lines of file till certain line (MOTIF + [num]) +''' +The script has to functions: + 1. Writing lines of file till certain line (MOTIF + [num]) + 2. Converting GLAM2 output to minimal meme-format +@params meme STING Path to 'meme' file generated from Meme suite +@parmas output STING Output file +@params num INT Number of motifs parsed from file + +@author René Wiegandt +@contact rene.wiegandt(at)mpi-bn.mpg.de +''' def main(): + args = parse_arguments() out = open(args.output, "w+") + + ''' + Create pattern where script should stop writing + For Example: + If num == 3, which means that you want the first/best 3 Motifs, the script + should stop writing lines to output if loop reaches line 'MOTIF 4' + ''' number = int(args.num) + 1 - motif = "MOTIF " + str(number) + break_header = "MOTIF " + str(number) + + # Pattern for motif header + pattern = re.compile("^MOTIF\s{2}(\d)+") + # Init count + count = 0 + with open(args.meme) as f: for line in f: - if motif in line: + ## do not write [count] lines after each header -> needed for meme-format + if count > 0: + count-=1 + continue + if pattern.match(line): + # if line is a motif header + count = 2 + ## + + if break_header in line: + # line matches breaking_header, e.g. 'MOTIF 4' break - out.write(line) + else: + out.write(line) if __name__ == "__main__": import argparse + import re main() From 88fa2980f9b7345bf02c65d0539c88f019dc7a42 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Wed, 19 Dec 2018 15:03:38 +0100 Subject: [PATCH 10/33] check whether jellyfish is installed --- bin/reduce_sequence.R | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/bin/reduce_sequence.R b/bin/reduce_sequence.R index 69fc764..d13f899 100644 --- a/bin/reduce_sequence.R +++ b/bin/reduce_sequence.R @@ -33,8 +33,8 @@ opt <- parse_args(opt_parser) #' @param motif_occurence Define how many motifs are expected per sequence. This value is used during kmer cutoff calculation. Default = 1 meaning that there should be approximately one motif per sequence. #' reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed", threads = NULL, clean = TRUE, minoverlap_kmer = kmer - 1, minoverlap_motif = ceiling(motif / 2), min_seq_length = max(c(motif, kmer)), motif_occurence = 1) { - if (system("jellyfish count --version") != 0) { - stop("Required programm jellyfish not found! Please check whether it is installed.") + if (system("which jellyfish", ignore.stdout = TRUE) != 0) { + stop("Required program jellyfish not found! Please check whether it is installed.") } if (missing(input)) { From e17d1dbbe25b8604e2ab5ffe1c18ee8d1c0a81a9 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Wed, 19 Dec 2018 15:03:56 +0100 Subject: [PATCH 11/33] check whether cdhit is installed --- bin/cdhit_wrapper.R | 4 ++++ 1 file changed, 4 insertions(+) diff --git a/bin/cdhit_wrapper.R b/bin/cdhit_wrapper.R index e5ed3e4..cf53ffc 100644 --- a/bin/cdhit_wrapper.R +++ b/bin/cdhit_wrapper.R @@ -70,6 +70,10 @@ opt <- parse_args(opt_parser) #' #' TODO check whether cdhit is installed cdhitest <- function(input, identity = 0.8, coverage = 8, output = "cluster.bed", clean = TRUE, threads = 1, global = 0, band_width = 20, memory = 800, word_length = 3, throw_away_sequences = 5, length_dif_cutoff_shorter_p = 0, length_dif_cutoff_shorter_n = 999999, alignment_coverage_longer_p = 0, alignment_coverage_longer_n = 99999999, alignment_coverage_shorter_p = 0, alignment_coverage_shorter_n = 99999999, max_unmatched_longer_p = 1, max_unmatched_shorter_p = 1, max_unmatched_both_n = 99999999, fast_cluster = 1, strand = 0, match = 2, mismatch = -2, gap = -6, gap_ext = -1, sort_cluster_by_size = 1) { + if (system("which cd-hit-est", ignore.stdout = FALSE) != 0) { + stop("Required program CD-HIT not found! Please check whether it is installed.") + } + if (missing(input)) { stop("Input parameter missing!") } From dcd185e80177a6fc873aaaf9c0347f1e7c68769c Mon Sep 17 00:00:00 2001 From: Schultheis Date: Wed, 19 Dec 2018 16:20:54 +0100 Subject: [PATCH 12/33] omit TODO --- bin/cdhit_wrapper.R | 1 - 1 file changed, 1 deletion(-) diff --git a/bin/cdhit_wrapper.R b/bin/cdhit_wrapper.R index cf53ffc..71bf881 100644 --- a/bin/cdhit_wrapper.R +++ b/bin/cdhit_wrapper.R @@ -68,7 +68,6 @@ opt <- parse_args(opt_parser) #' @param gat_ext Gap extension score. Default = -1 (CD-HIT parameter) #' @param sort_cluster_by_size Either sort cluster by decreasing length (= 0) or by decreasing size (= 1). Default = 1 (CD-HIT parameter) #' -#' TODO check whether cdhit is installed cdhitest <- function(input, identity = 0.8, coverage = 8, output = "cluster.bed", clean = TRUE, threads = 1, global = 0, band_width = 20, memory = 800, word_length = 3, throw_away_sequences = 5, length_dif_cutoff_shorter_p = 0, length_dif_cutoff_shorter_n = 999999, alignment_coverage_longer_p = 0, alignment_coverage_longer_n = 99999999, alignment_coverage_shorter_p = 0, alignment_coverage_shorter_n = 99999999, max_unmatched_longer_p = 1, max_unmatched_shorter_p = 1, max_unmatched_both_n = 99999999, fast_cluster = 1, strand = 0, match = 2, mismatch = -2, gap = -6, gap_ext = -1, sort_cluster_by_size = 1) { if (system("which cd-hit-est", ignore.stdout = FALSE) != 0) { stop("Required program CD-HIT not found! Please check whether it is installed.") From 4c16f6f796238619700711f5030c5ae983e8e74a Mon Sep 17 00:00:00 2001 From: Schultheis Date: Wed, 19 Dec 2018 16:55:13 +0100 Subject: [PATCH 13/33] check for header and forward it if provided --- bin/cdhit_wrapper.R | 19 +++++++++++++------ 1 file changed, 13 insertions(+), 6 deletions(-) diff --git a/bin/cdhit_wrapper.R b/bin/cdhit_wrapper.R index 71bf881..c5f7230 100644 --- a/bin/cdhit_wrapper.R +++ b/bin/cdhit_wrapper.R @@ -68,6 +68,8 @@ opt <- parse_args(opt_parser) #' @param gat_ext Gap extension score. Default = -1 (CD-HIT parameter) #' @param sort_cluster_by_size Either sort cluster by decreasing length (= 0) or by decreasing size (= 1). Default = 1 (CD-HIT parameter) #' +#' @details If there is a header supplied other then the default data.table nameing scheme ('V1', 'V2', etc.) it will be kept and extended. +#' cdhitest <- function(input, identity = 0.8, coverage = 8, output = "cluster.bed", clean = TRUE, threads = 1, global = 0, band_width = 20, memory = 800, word_length = 3, throw_away_sequences = 5, length_dif_cutoff_shorter_p = 0, length_dif_cutoff_shorter_n = 999999, alignment_coverage_longer_p = 0, alignment_coverage_longer_n = 99999999, alignment_coverage_shorter_p = 0, alignment_coverage_shorter_n = 99999999, max_unmatched_longer_p = 1, max_unmatched_shorter_p = 1, max_unmatched_both_n = 99999999, fast_cluster = 1, strand = 0, match = 2, mismatch = -2, gap = -6, gap_ext = -1, sort_cluster_by_size = 1) { if (system("which cd-hit-est", ignore.stdout = FALSE) != 0) { stop("Required program CD-HIT not found! Please check whether it is installed.") @@ -80,15 +82,19 @@ cdhitest <- function(input, identity = 0.8, coverage = 8, output = "cluster.bed" message("Loading bed.") # load bed if neccessary if (!data.table::is.data.table(input)) { - bed_table <- data.table::fread(input = input, header = FALSE) + bed_table <- data.table::fread(input = input) } else { bed_table <- input } + # check if there are column names to keep them + default_col_names <- grepl(pattern = "^V+\\d$", names(bed_table), perl = TRUE) + keep_col_names <- ifelse(any(default_col_names), FALSE, TRUE) + # check for duplicated names - if (anyDuplicated(bed_table[, "V4"])) { + if (anyDuplicated(bed_table[, 4])) { warning("Found duplicated names. Making names unique.") - bed_table[, V4 := make.unique(V4)] + bed_table[, 4 := make.unique(names(bed_table)[4])] } message("Convert to fasta.") @@ -140,17 +146,18 @@ cdhitest <- function(input, identity = 0.8, coverage = 8, output = "cluster.bed" system(command = cluster_call, wait = TRUE) # load reformated file - cluster <- data.table::fread(cluster_file) + cluster <- data.table::fread(cluster_file)[, c("id", "clstr")] + names(cluster) <- c("id", "cluster") ### add cluster to bed_table - result <- merge(x = bed_table, y = cluster[, c("id", "clstr")], by.x = "V4", by.y = "id", sort = FALSE)[, union(names(bed_table), names(cluster)[2]), with = FALSE] + result <- merge(x = bed_table, y = cluster, by.x = names(bed_table)[4], by.y = "id", sort = FALSE)[, union(names(bed_table), names(cluster)[2]), with = FALSE] # delete files if (clean) { file.remove(fasta_file, paste0(cdhit_output, ".clstr"), cdhit_output, cluster_file) } - data.table::fwrite(x = result, file = output, sep = "\t", col.names = FALSE) + data.table::fwrite(x = result, file = output, sep = "\t", col.names = keep_col_names) } # call function with given parameter if not in interactive context (e.g. run from shell) From 5a7c84eb969aed0d111ba320a0b15310c275508d Mon Sep 17 00:00:00 2001 From: Schultheis Date: Wed, 19 Dec 2018 17:11:46 +0100 Subject: [PATCH 14/33] automatically detect and keep column names if provided --- bin/reduce_sequence.R | 27 ++++++++++++++++++++++----- 1 file changed, 22 insertions(+), 5 deletions(-) diff --git a/bin/reduce_sequence.R b/bin/reduce_sequence.R index d13f899..f9e8340 100644 --- a/bin/reduce_sequence.R +++ b/bin/reduce_sequence.R @@ -32,6 +32,8 @@ opt <- parse_args(opt_parser) #' @param min_seq_length Remove sequences below this length. Defaults to the maximum value of motif and kmer and can not be lower. #' @param motif_occurence Define how many motifs are expected per sequence. This value is used during kmer cutoff calculation. Default = 1 meaning that there should be approximately one motif per sequence. #' +#' @details If there is a header supplied other then the default data.table nameing scheme ('V1', 'V2', etc.) it will be kept. +#' reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed", threads = NULL, clean = TRUE, minoverlap_kmer = kmer - 1, minoverlap_motif = ceiling(motif / 2), min_seq_length = max(c(motif, kmer)), motif_occurence = 1) { if (system("which jellyfish", ignore.stdout = TRUE) != 0) { stop("Required program jellyfish not found! Please check whether it is installed.") @@ -49,7 +51,17 @@ reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed" message("Loading bed...") # load bed # columns: chr, start, end, name, ..., sequence - bed_table <- data.table::fread(input = input, header = FALSE) + bed_table <- data.table::fread(input = input) + + # check for header and save it if provided + default_col_names <- grepl(pattern = "^V+\\d$", names(bed_table), perl = TRUE) + if (!any(default_col_names)) { + keep_col_names <- TRUE + col_names <- names(bed_table) + } else { + keep_col_names <- FALSE + } + names(bed_table)[1:4] <- c("chr", "start", "end", "name") names(bed_table)[ncol(bed_table)] <- "sequence" # index @@ -123,18 +135,23 @@ reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed" merged[, sequence := stringr::str_sub(sequence, relative_start, relative_end)] # bed files count from 0 - merged[, data.table::`:=`(relative_start = relative_start - 1, relative_end = relative_end - 1)] + merged[, `:=`(relative_start = relative_start - 1, relative_end = relative_end - 1)] # change start end location - merged[, data.table::`:=`(start = start + relative_start, end = start + relative_end)] + merged[, `:=`(start = start + relative_start, end = start + relative_end)] # clean table - merged[, data.table::`:=`(relative_start = NULL, relative_end = NULL, width = NULL)] + merged[, `:=`(relative_start = NULL, relative_end = NULL, width = NULL)] if (clean) { file.remove(fasta_file, count_output_binary, mer_count_table) } - data.table::fwrite(merged, file = output, sep = "\t", col.names = FALSE) + # keep provided column names + if (keep_col_names) { + names(merged) <- col_names + } + + data.table::fwrite(merged, file = output, sep = "\t", col.names = keep_col_names) } #' Predict how many interesting kmer are possible for the given data. From 97464ca036c2f0830af7b2497e3c7d8f72e94e47 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Wed, 19 Dec 2018 17:17:12 +0100 Subject: [PATCH 15/33] added author; better missing input error --- bin/cdhit_wrapper.R | 5 +++-- 1 file changed, 3 insertions(+), 2 deletions(-) diff --git a/bin/cdhit_wrapper.R b/bin/cdhit_wrapper.R index c5f7230..fcbd5eb 100644 --- a/bin/cdhit_wrapper.R +++ b/bin/cdhit_wrapper.R @@ -33,7 +33,8 @@ option_list <- list( ) opt_parser <- OptionParser(option_list = option_list, - description = "CD-HIT-EST Wrapper function. See https://github.com/weizhongli/cdhit/wiki for more information regarding CD-HITs parameters.") + description = "CD-HIT-EST Wrapper function. See https://github.com/weizhongli/cdhit/wiki for more information regarding CD-HITs parameters.", + epilogue = "Author: Hendrik Schultheis ") opt <- parse_args(opt_parser) @@ -76,7 +77,7 @@ cdhitest <- function(input, identity = 0.8, coverage = 8, output = "cluster.bed" } if (missing(input)) { - stop("Input parameter missing!") + stop("No input specified! Please forward a valid bed-file.") } message("Loading bed.") From cc532bf00dc6d0a967e0fced688757f7f67a0fb1 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Wed, 19 Dec 2018 17:17:29 +0100 Subject: [PATCH 16/33] added author --- bin/reduce_sequence.R | 3 ++- 1 file changed, 2 insertions(+), 1 deletion(-) diff --git a/bin/reduce_sequence.R b/bin/reduce_sequence.R index f9e8340..6df5bdc 100644 --- a/bin/reduce_sequence.R +++ b/bin/reduce_sequence.R @@ -15,7 +15,8 @@ option_list <- list( ) opt_parser <- OptionParser(option_list = option_list, - description = "Reduces each sequence to its most frequent region.") + description = "Reduces each sequence to its most frequent region.", + epilogue = "Author: Hendrik Schultheis ") opt <- parse_args(opt_parser) From 8389226abaf3c6724b2978fc38bc08c9d06196b3 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ren=C3=A9=20Wiegandt?= Date: Wed, 19 Dec 2018 15:34:43 -0500 Subject: [PATCH 17/33] Fixed typos in get_best_motif.py --- bin/get_best_motif.py | 8 ++++---- 1 file changed, 4 insertions(+), 4 deletions(-) diff --git a/bin/get_best_motif.py b/bin/get_best_motif.py index 37eccc5..a506bd8 100644 --- a/bin/get_best_motif.py +++ b/bin/get_best_motif.py @@ -11,11 +11,11 @@ def parse_arguments(): return args ''' -The script has to functions: +The script has two functions: 1. Writing lines of file till certain line (MOTIF + [num]) 2. Converting GLAM2 output to minimal meme-format -@params meme STING Path to 'meme' file generated from Meme suite -@parmas output STING Output file +@params meme STRING Path to 'meme' file generated from Meme suite +@parmas output STRING Output file @params num INT Number of motifs parsed from file @author René Wiegandt @@ -25,7 +25,7 @@ def main(): args = parse_arguments() out = open(args.output, "w+") - + ''' Create pattern where script should stop writing For Example: From 2fca1581a160844d89be2fda2aea322a1740b9b3 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ren=C3=A9=20Wiegandt?= Date: Wed, 19 Dec 2018 15:35:32 -0500 Subject: [PATCH 18/33] Reads BED-files with or without header --- bin/bed_to_fasta.R | 116 ++++++++++++++++++++++----------------------- 1 file changed, 58 insertions(+), 58 deletions(-) diff --git a/bin/bed_to_fasta.R b/bin/bed_to_fasta.R index bb4d3dc..73ea732 100644 --- a/bin/bed_to_fasta.R +++ b/bin/bed_to_fasta.R @@ -1,58 +1,58 @@ -#!/usr/bin/env Rscript -library("optparse") - -option_list <- list( - make_option(opt_str = c("-i", "--input"), default = NULL, help = "Input bed-file. Second last column must be sequences and last column must be the cluster_id.", metavar = "character"), - make_option(opt_str = c("-p", "--prefix"), default = "" , help = "Prefix for file names. Default = '%default'", metavar = "character"), - make_option(opt_str = c("-m", "--min_seq"), default = 100, help = "Minimum amount of sequences in clusters. Default = %default", metavar = "integer") -) - -opt_parser <- OptionParser(option_list = option_list, - description = "Convert BED-file to one FASTA-file per cluster") - -opt <- parse_args(opt_parser) - -#' Splitting BED-files depending on their cluster. -#' The Sequences of each cluster are writen as an FASTA-file. -#' @param bedInput BED-file with sequences and cluster-id as last two columns: -#' Sequence: second last column; Cluster ID: last column -#' @param prefix prefix for filenames -#' @param min_seq min. number of sequences per cluster -#' -#' @author René Wiegandt -#' @contact rene.wiegandt(at)mpi-bn.mpg.de -bed_to_fasta <- function(bedInput, prefix = "", min_seq = 100){ - - if(is.null(bedInput)){ - stop("ERROR: Input parameter cannot be null! Please specify the input parameter.") - } - - bed <- data.table::fread(bedInput, header = FALSE, sep = "\t") - - # Get last column of data.table, which refers to the cluster, as a vector. - cluster_no <- as.vector(bed[[ncol(bed)]]) - - # Split data.table bed on its last column (cluster_no) into list of data.frames - clusters <- split(bed, cluster_no, sorted = TRUE, flatten = FALSE) - - # For each data.frame(cluster) in list clusters: - discard <- lapply(1:length(clusters), function(i){ - clust <- as.data.frame(clusters[i]) - # Filter data.tables(clusters), which are to small - if (nrow(clust) >= as.numeric(min_seq) ) { - # Get second last column, which contains the nucleotide sequences - sequences <- as.list(clust[[ncol(clust) - 1]]) - # Create filename - outfile <- paste0(prefix,"_cluster_",i - 1,".FASTA") - # Write fasta file - seqinr::write.fasta(sequences = sequences, names = clust[[4]], file.out = outfile, as.string = TRUE) - } else { - print(paste0("Cluster: ",i," is to small")) - } - }) -} - -# run function bed_to_fasta with given parameteres if not in interactive context (e.g. run from shell) -if (!interactive()) { - bed_to_fasta(opt$input, opt$prefix, opt$min_seq) -} +#!/usr/bin/env Rscript +library("optparse") + +option_list <- list( + make_option(opt_str = c("-i", "--input"), default = NULL, help = "Input bed-file. Second last column must be sequences and last column must be the cluster_id.", metavar = "character"), + make_option(opt_str = c("-p", "--prefix"), default = "" , help = "Prefix for file names. Default = '%default'", metavar = "character"), + make_option(opt_str = c("-m", "--min_seq"), default = 100, help = "Minimum amount of sequences in clusters. Default = %default", metavar = "integer") +) + +opt_parser <- OptionParser(option_list = option_list, + description = "Convert BED-file to one FASTA-file per cluster") + +opt <- parse_args(opt_parser) + +#' Splitting BED-files depending on their cluster. +#' The Sequences of each cluster are writen as an FASTA-file. +#' @param bedInput BED-file with sequences and cluster-id as last two columns: +#' Sequence: second last column; Cluster ID: last column +#' @param prefix prefix for filenames +#' @param min_seq min. number of sequences per cluster +#' +#' @author René Wiegandt +#' @contact rene.wiegandt(at)mpi-bn.mpg.de +bed_to_fasta <- function(bedInput, prefix = "", min_seq = 100){ + + if (is.null(bedInput)) { + stop("ERROR: Input parameter cannot be null! Please specify the input parameter.") + } + + bed <- data.table::fread(bedInput, sep = "\t") + + # Get last column of data.table, which refers to the cluster, as a vector. + cluster_no <- as.vector(bed[[ncol(bed)]]) + + # Split data.table bed on its last column (cluster_no) into list of data.frames + clusters <- split(bed, cluster_no, sorted = TRUE, flatten = FALSE) + + # For each data.frame(cluster) in list clusters: + discard <- lapply(1:length(clusters), function(i){ + clust <- as.data.frame(clusters[i]) + # Filter data.tables(clusters), which are to small + if (nrow(clust) >= as.numeric(min_seq) ) { + # Get second last column, which contains the nucleotide sequences + sequences <- as.list(clust[[ncol(clust) - 1]]) + # Create filename + outfile <- paste0(prefix,"_cluster_",i - 1,".FASTA") + # Write fasta file + seqinr::write.fasta(sequences = sequences, names = clust[[4]], file.out = outfile, as.string = TRUE) + } else { + print(paste0("Cluster: ",i," is to small")) + } + }) +} + +# run function bed_to_fasta with given parameteres if not in interactive context (e.g. run from shell) +if (!interactive()) { + bed_to_fasta(opt$input, opt$prefix, opt$min_seq) +} From 4dea8e413781953175fdef9679a408058bd27269 Mon Sep 17 00:00:00 2001 From: renewiegandt Date: Thu, 20 Dec 2018 11:51:52 +0100 Subject: [PATCH 19/33] Imporved description for installation in README.md --- README.md | 50 +++++++++++++++++++++++--------------------------- 1 file changed, 23 insertions(+), 27 deletions(-) diff --git a/README.md b/README.md index 1aa01e6..41ee001 100644 --- a/README.md +++ b/README.md @@ -7,38 +7,16 @@ For further information read the [documentation](https://github.molgen.mpg.de/lo ## Dependencies * [conda](https://conda.io/docs/user-guide/install/linux.html) * [Nextflow](https://www.nextflow.io/) -* [MEME-Suite](http://meme-suite.org/doc/install.html?man_type=web) ## Installation -Start with installing all dependencies listed above. It is required to set the [enviroment paths for meme-suite](http://meme-suite.org/doc/install.html?man_type=web#installingtar). -this can be done with following commands: -``` -export PATH=[meme-suite instalation path]/libexec/meme-[meme-suite version]:$PATH -export PATH=[meme-suite instalation path]/bin:$PATH -``` - +Start with installing all dependencies listed above (Nextflow, conda) and downloading all files from the [GitHub repository](https://github.molgen.mpg.de/loosolab/masterJLU2018). -Download all files from the [GitHub repository](https://github.molgen.mpg.de/loosolab/masterJLU2018). -The Nextflow-script needs a conda enviroment to run. Nextflow can create the needed enviroment from the given yaml-file. -On some systems Nextflow exits the run with following error: -``` -Caused by: - Failed to create Conda environment - command: conda env create --prefix --file env.yml - status : 143 - message: -``` -If this error occurs you have to create the enviroment before starting the pipeline. -To create this enviroment you need the yml-file from the repository. -Run the following commands to create the enviroment: -```console -path=[Path to given masterenv.yml file] -conda env create --name masterenv -f=$path -``` -When the enviroment is created, set the variable 'path_env' in the configuration file as the path to it. +Every other dependency will be automatically installed by Nextflow using conda. For that a new conda enviroment will be created, which can be found in the from Nextflow created work directory after the first pipeline run. +It is **not** required to create and activate the enviroment from the yaml-file beforehand. **Important Note:** For conda the channel bioconda needs to be set as highest priority! This is required due to two differnt packages with the same name in different channels. For the pipeline the package jellyfish from the channel bioconda is needed and **NOT** the jellyfisch package from the channel conda-forge! + ## Quick Start ```console nextflow run pipeline.nf --bigwig [BigWig-file] --bed [BED-file] --genome_fasta [FASTA-file] --motif_db [MEME-file] --config [UROPA-config-file] @@ -105,6 +83,24 @@ Optional arguments: All arguments can be set in the configuration files ``` +For further information read the [documentation](https://github.molgen.mpg.de/loosolab/masterJLU2018/wiki) +## Known issues +The Nextflow-script needs a conda enviroment to run. Nextflow creates the needed enviroment from the given yaml-file. +On some systems Nextflow exits the run with following error: +``` +Caused by: + Failed to create Conda environment + command: conda env create --prefix --file env.yml + status : 143 + message: +``` +If this error occurs you have to create the enviroment before starting the pipeline. +To create this enviroment you need the yml-file from the repository. +Run the following commands to create the enviroment: +```console +path=[Path to given masterenv.yml file] +conda env create --name masterenv -f $path +``` +When the enviroment is created, set the variable 'path_env' in the configuration file as the path to it. -For further information read the [documentation](https://github.molgen.mpg.de/loosolab/masterJLU2018/wiki) From 1a7a812b3f667a710438e05cac3d9f2b91ba4983 Mon Sep 17 00:00:00 2001 From: renewiegandt Date: Thu, 20 Dec 2018 11:58:11 +0100 Subject: [PATCH 20/33] Removed snakemake from yaml-file --- masterenv.yml | 1 - 1 file changed, 1 deletion(-) diff --git a/masterenv.yml b/masterenv.yml index 211f4e1..a2f13d6 100644 --- a/masterenv.yml +++ b/masterenv.yml @@ -14,7 +14,6 @@ dependencies: - r-stringr - r-optparse - bioconductor-iranges - - snakemake - meme - moods - biopython From 4844609d709c67e353c4727d0247ee0287bedf55 Mon Sep 17 00:00:00 2001 From: renewiegandt Date: Thu, 20 Dec 2018 13:14:41 +0100 Subject: [PATCH 21/33] Set parameter organism as required wihtout an default value --- pipeline.nf | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/pipeline.nf b/pipeline.nf index 55b7c26..3d51f1b 100644 --- a/pipeline.nf +++ b/pipeline.nf @@ -55,10 +55,10 @@ params.best_motif = 3 // Top n motifs per cluster //creating_gtf - params.organism="hg38" + params.organism="" params.tissue="" -if (params.bigwig == "" || params.bed == "" || params.genome_fasta == "" || params.motif_db == "" || params.config == "" || "${params.help}" != "0"){ +if (params.bigwig == "" || params.bed == "" || params.organism == "" || params.genome_fasta == "" || params.motif_db == "" || params.config == "" || "${params.help}" != "0"){ log.info """ Usage: nextflow run pipeline.nf --bigwig [BigWig-file] --bed [BED-file] --genome_fasta [FASTA-file] --motif_db [MEME-file] --config [UROPA-config-file] @@ -70,6 +70,7 @@ Required arguments: --config Path to UROPA configuration file --create_known_tfbs_path Path to directory where output from tfbsscan (known motifs) are stored. Path can be set as tfbs_path in next run. (Default: './') + --organism Input organism [hg38 | hg19 | mm9 | mm10] --out Output Directory (Default: './out/') Optional arguments: @@ -115,7 +116,6 @@ Optional arguments: --motif_similarity_thresh FLOAT Threshold for motif similarity score (Default: 0.00001) Creating GTF: - --organism [hg38 | hg19 | mm9 | mm10] Input organism --tissues List/String List of one or more keywords for tissue-/category-activity, categories must be specified as in JSON config All arguments can be set in the configuration files From d60faa704e974a4b66af3bfb582268e4817397d7 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Thu, 20 Dec 2018 13:21:35 +0100 Subject: [PATCH 22/33] spell check --- bin/cdhit_wrapper.R | 20 ++++++++++---------- 1 file changed, 10 insertions(+), 10 deletions(-) diff --git a/bin/cdhit_wrapper.R b/bin/cdhit_wrapper.R index fcbd5eb..38b863a 100644 --- a/bin/cdhit_wrapper.R +++ b/bin/cdhit_wrapper.R @@ -4,7 +4,7 @@ library("optparse") option_list <- list( make_option(opt_str = c("-i", "--input"), default = NULL, help = "Input bed-file. Fourth column is expected to contain names, last column must be sequences.", metavar = "character"), make_option(opt_str = c("-c", "--identity"), default = 0.8, help = "Identity threshold. Default = %default (CD-HIT parameter)", metavar = "double >= 0.8"), - make_option(opt_str = c("-A", "--coverage"), default = 8, help = "Minimal alignment length for both sequences in nucelotides. Default = %default (CD-HIT parameter)", metavar = "integer"), + make_option(opt_str = c("-A", "--coverage"), default = 8, help = "Minimal alignment length for both sequences in nucleotides. Default = %default (CD-HIT parameter)", metavar = "integer"), make_option(opt_str = c("-o", "--output"), default = "cluster.bed", help = "Output file same as input but with appended column of cluster numbers. Default = %default", metavar = "character"), make_option(opt_str = c("--clean"), default = TRUE, help = "Delete all temporary files. Default = %default", metavar = "logical"), make_option(opt_str = c("-G", "--global"), default = 0, help = "Global sequence identity = 1, local = 0. Default = %default (CD-HIT parameter)", metavar = "integer"), @@ -14,7 +14,7 @@ option_list <- list( make_option(opt_str = c("-n", "--word_length"), default = 3, help = "Default = %default (CD-HIT parameter)", metavar = "integer"), make_option(opt_str = c("-l", "--throw_away_sequences"), default = 5, help = "Maximum length of sequences thrown away. Default = %default (CD-HIT parameter)", metavar = "integer"), # make_option(opt_str = c("-d", "--description")), # can not produce bed if this is != 0 - make_option(opt_str = c("-s", "--length_dif_cutoff_shorter_p"), default = 0.0, help = "Shorter sequences length must be at least x percent of longer sequenecs length. Default = %default (CD-HIT parameter)", metavar = "double"), + make_option(opt_str = c("-s", "--length_dif_cutoff_shorter_p"), default = 0.0, help = "Shorter sequences length must be at least x percent of longer sequences length. Default = %default (CD-HIT parameter)", metavar = "double"), make_option(opt_str = c("-S", "--length_dif_cutoff_shorter_n"), default = 999999, help = "Length difference between sequences can not be higher than x nucleotides. Default = %default (CD-HIT parameter)", metavar = "integer"), make_option(opt_str = c("-e", "--alignment_coverage_longer_p"), default = 0.0, help = "Alignment must cover x percent of longer sequence. Default = %default (CD-HIT parameter: -aL)", metavar = "double"), make_option(opt_str = c("-E", "--alignment_coverage_longer_n"), default = 99999999, help = "There can not be more than x unaligned nucleotides on the longer sequence. Default = %default (CD-HIT parameter: -AL)", metavar = "integer"), @@ -22,7 +22,7 @@ option_list <- list( make_option(opt_str = c("-F", "--alignment_coverage_shorter_n"), default = 99999999, help = "There can not be more than x unaligned nucleotides on the longer sequence. Default = %default (CD-HIT parameter: -AS)", metavar = "integer"), make_option(opt_str = c("-w", "--max_unmatched_longer_p"), default = 1.0, help = "Maximum unmatched percentage on longer sequence (excludes leading tailing gap). Default = %default (CD-HIT parameter: -uL)", metavar = "double"), make_option(opt_str = c("-W", "--max_unmatched_shorter_p"), default = 1.0, help = "Maximum unmatched percentage on shorter sequence (excludes leading tailing gap). Default = %default (CD-HIT parameter: -uS)", metavar = "double"), - make_option(opt_str = c("-U", "--max_unmatched_both_n"), default = 99999999, help = "Maximum unmatched nucleotied on both sequences (excludes leading tailing gap). Default = %default (CD-HIT parameter)", metavar = "integer"), + make_option(opt_str = c("-U", "--max_unmatched_both_n"), default = 99999999, help = "Maximum unmatched nucleotide on both sequences (excludes leading tailing gap). Default = %default (CD-HIT parameter)", metavar = "integer"), make_option(opt_str = c("-g", "--fast_cluster"), default = 1, help = "Cluster sequence in first cluster that meets threshold = 0 (fast). Or cluster in best Cluster = 1 (accurate). Default = %default (CD-HIT parameter)", metavar = "integer"), make_option(opt_str = c("-r", "--strand"), default = 0, help = "Align +/+ & +/- (= 1). Or align only +/+ (= 0). Default = %default (CD-HIT parameter)", metavar = "integer"), make_option(opt_str = c("--match"), default = 2, help = "Matching score. Default = %default (CD-HIT parameter)", metavar = "integer"), @@ -43,8 +43,8 @@ opt <- parse_args(opt_parser) #' #' @param input Data.table or file in bed format (requires names in fourth column and sequences in last column). #' @param identity Identity threshold. Default = 0.8 (CD-HIT parameter) -#' @param coverage Minimal alignment length for both sequences in nucelotides. Default = 8 (CD-HIT parameter) -#' @param output Clustered bedfile. Adds cluster number in last column (numbering depend on sort_by_cluster_size parameter). Default = cluster.bed +#' @param coverage Minimal alignment length for both sequences in nucleotides. Default = 8 (CD-HIT parameter) +#' @param output Clustered bed-file. Adds cluster number in last column (numbering depend on sort_by_cluster_size parameter). Default = cluster.bed #' @param clean Clean up after run. Default = TRUE #' @param threads Number of threads to use (0 = all cores). Default = 1 (CD-HIT parameter) #' @param global Global sequence identity = 1, local = 0. Default = 0 (CD-HIT parameter) @@ -52,7 +52,7 @@ opt <- parse_args(opt_parser) #' @param memory Memory limit in MB. 0 for unlimited. Default = 800 (CD-HIT parameter) #' @param word_length Default = 3 (CD-HIT parameter) #' @param throw_away_sequences Maximum length of sequences thrown away. Default = %default (CD-HIT parameter) -#' @param length_dif_cutoff_shorter_p Shorter sequences length must be at least x percent of longer sequenecs length. Default = 0 (CD-HIT parameter) +#' @param length_dif_cutoff_shorter_p Shorter sequences length must be at least x percent of longer sequences length. Default = 0 (CD-HIT parameter) #' @param length_dif_cutoff_shorter_n Length difference between sequences can not be higher than x nucleotides. Default = 999999 (CD-HIT parameter) #' @param alignment_coverage_longer_p Alignment must cover x percent of longer sequence. Default = 0 (CD-HIT parameter) #' @param alignment_coverage_longer_n There can not be more than x unaligned nucleotides on the longer sequence. Default = 99999999 (CD-HIT parameter) @@ -60,16 +60,16 @@ opt <- parse_args(opt_parser) #' @param alignment_coverage_shorter_n There can not be more than x unaligned nucleotides on the longer sequence. Default = 99999999 (CD-HIT parameter) #' @param max_unmatched_longer_p Maximum unmatched percentage on longer sequence (excludes leading tailing gap). Default = 1 (CD-HIT parameter) #' @param max_unmatched_shorter_p Maximum unmatched percentage on shorter sequence (excludes leading tailing gap). Default = 1 (CD-HIT parameter) -#' @param max_unmatched_both_n Maximum unmatched nucleotied on both sequences (excludes leading tailing gap). Default = 99999999 (CD-HIT parameter) +#' @param max_unmatched_both_n Maximum unmatched nucleotide on both sequences (excludes leading tailing gap). Default = 99999999 (CD-HIT parameter) #' @param fast_cluster Cluster sequence in first cluster that meets threshold = 0 (fast). Or cluster in best Cluster = 1 (accurate). Default = 1 (CD-HIT parameter) #' @param strand Align +/+ & +/- (= 1). Or align only +/+ (= 0). Default = 0 (CD-HIT parameter) #' @param match Matching score. Default = 2 (CD-HIT parameter) #' @param mismatch Mismatch score. Default = -2 (CD-HIT parameter) #' @param gap Gap score. Default = -6 (CD-HIT parameter) -#' @param gat_ext Gap extension score. Default = -1 (CD-HIT parameter) +#' @param gap_ext Gap extension score. Default = -1 (CD-HIT parameter) #' @param sort_cluster_by_size Either sort cluster by decreasing length (= 0) or by decreasing size (= 1). Default = 1 (CD-HIT parameter) #' -#' @details If there is a header supplied other then the default data.table nameing scheme ('V1', 'V2', etc.) it will be kept and extended. +#' @details If there is a header supplied other then the default data.table naming scheme ('V1', 'V2', etc.) it will be kept and extended. #' cdhitest <- function(input, identity = 0.8, coverage = 8, output = "cluster.bed", clean = TRUE, threads = 1, global = 0, band_width = 20, memory = 800, word_length = 3, throw_away_sequences = 5, length_dif_cutoff_shorter_p = 0, length_dif_cutoff_shorter_n = 999999, alignment_coverage_longer_p = 0, alignment_coverage_longer_n = 99999999, alignment_coverage_shorter_p = 0, alignment_coverage_shorter_n = 99999999, max_unmatched_longer_p = 1, max_unmatched_shorter_p = 1, max_unmatched_both_n = 99999999, fast_cluster = 1, strand = 0, match = 2, mismatch = -2, gap = -6, gap_ext = -1, sort_cluster_by_size = 1) { if (system("which cd-hit-est", ignore.stdout = FALSE) != 0) { @@ -81,7 +81,7 @@ cdhitest <- function(input, identity = 0.8, coverage = 8, output = "cluster.bed" } message("Loading bed.") - # load bed if neccessary + # load bed if necessary if (!data.table::is.data.table(input)) { bed_table <- data.table::fread(input = input) } else { From 46cfc59119899bb65efadcc007e924707b97cb75 Mon Sep 17 00:00:00 2001 From: renewiegandt Date: Thu, 20 Dec 2018 13:39:13 +0100 Subject: [PATCH 23/33] Added Parameter gtf_path. If path is set process create_gtf will be skipped --- pipeline.nf | 17 ++++++++++++++--- 1 file changed, 14 insertions(+), 3 deletions(-) diff --git a/pipeline.nf b/pipeline.nf index 3d51f1b..1be18be 100644 --- a/pipeline.nf +++ b/pipeline.nf @@ -9,6 +9,7 @@ params.tfbs_path="" params.create_known_tfbs_path = "./" params.help = 0 + params.get_path="" params.out = "./out/" //peak_calling @@ -68,8 +69,6 @@ Required arguments: --genome_fasta Path to genome in FASTA-format --motif_db Path to motif-database in MEME-format --config Path to UROPA configuration file - --create_known_tfbs_path Path to directory where output from tfbsscan (known motifs) are stored. - Path can be set as tfbs_path in next run. (Default: './') --organism Input organism [hg38 | hg19 | mm9 | mm10] --out Output Directory (Default: './out/') @@ -77,6 +76,9 @@ Optional arguments: --help [0|1] 1 to show this help message. (Default: 0) --tfbs_path Path to directory with output from tfbsscan. If given tfbsscan will not be run. + --create_known_tfbs_path Path to directory where output from tfbsscan (known motifs) are stored. + Path can be set as tfbs_path in next run. (Default: './') + --gtf_path Path to gtf-file. If path is set the process which creats a gtf-file is skipped. Footprint extraction: --window_length INT This parameter sets the length of a sliding window. (Default: 200) @@ -578,7 +580,10 @@ process create_GTF { publishDir "${params.out}/gtf/", mode: 'copy' output: - file ('*.gtf') into gtf_for_uropa + file ('*.gtf') into gtf + + when: + gtf_path == "" script: """ @@ -586,6 +591,12 @@ process create_GTF { """ } +if (gtf_path == "") { + gtf_for_uropa = gtf +} else { + gtf_for_uropa = Channel.fromPath(params.gtf_path) +} + /* bed_for_final_filter.combine(gtf_for_uropa).set {uropa_in} From 65076439ec5990c384b63fc9c9bc7781428bac86 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Thu, 20 Dec 2018 13:53:09 +0100 Subject: [PATCH 24/33] spell check --- bin/reduce_sequence.R | 76 +++++++++++++++++++++---------------------- 1 file changed, 38 insertions(+), 38 deletions(-) diff --git a/bin/reduce_sequence.R b/bin/reduce_sequence.R index 6df5bdc..cf91187 100644 --- a/bin/reduce_sequence.R +++ b/bin/reduce_sequence.R @@ -3,15 +3,15 @@ library("optparse") option_list <- list( make_option(opt_str = c("-i", "--input"), default = NULL, help = "Input bed-file. Last column must be sequences.", metavar = "character"), - make_option(opt_str = c("-k", "--kmer"), default = 10, help = "Kmer length. Default = %default", metavar = "integer"), + make_option(opt_str = c("-k", "--kmer"), default = 10, help = "k-mer length. Default = %default", metavar = "integer"), make_option(opt_str = c("-m", "--motif"), default = 10, help = "Estimated motif length. Default = %default", metavar = "integer"), make_option(opt_str = c("-o", "--output"), default = "reduced.bed", help = "Output file. Default = %default", metavar = "character"), make_option(opt_str = c("-t", "--threads"), default = 1, help = "Number of threads to use. Use 0 for all available cores. Default = %default", metavar = "integer"), make_option(opt_str = c("-c", "--clean"), default = TRUE, help = "Delete all temporary files. Default = %default", metavar = "logical"), - make_option(opt_str = c("-s", "--min_seq_length"), default = NULL, help = "Remove sequences below this length. Defaults to the maximum value of motif and kmer and can not be lower.", metavar = "integer", type = "integer"), - make_option(opt_str = c("-n", "--minoverlap_kmer"), default = NULL, help = "Minimum required overlap between kmer. Used to create reduced sequence ranges out of merged kmer. Can not be greater than kmer length. Default = kmer - 1", metavar = "integer", type = "integer"), - make_option(opt_str = c("-v", "--minoverlap_motif"), default = NULL, help = "Minimum required overlap between motif and kmer to consider kmer significant. Used for kmer cutoff calculation. Can not be greater than motif and kmer length. Default = ceiling(motif / 2)", metavar = "integer", type = "integer"), - make_option(opt_str = c("-f", "--motif_occurence"), default = 1, help = "Define how many motifs are expected per sequence. This value is used during kmer cutoff calculation. Default = %default meaning that there should be approximately one motif per sequence.", metavar = "double") + make_option(opt_str = c("-s", "--min_seq_length"), default = NULL, help = "Remove sequences below this length. Defaults to the maximum value of motif and k-mer and can not be lower.", metavar = "integer", type = "integer"), + make_option(opt_str = c("-n", "--minoverlap_kmer"), default = NULL, help = "Minimum required overlap between k-mer. Used to create reduced sequence ranges out of merged k-mer. Can not be greater than k-mer length. Default = kmer - 1", metavar = "integer", type = "integer"), + make_option(opt_str = c("-v", "--minoverlap_motif"), default = NULL, help = "Minimum required overlap between motif and k-mer to consider k-mer significant. Used for k-mer cutoff calculation. Can not be greater than motif and k-mer length. Default = ceiling(motif / 2)", metavar = "integer", type = "integer"), + make_option(opt_str = c("-f", "--motif_occurrence"), default = 1, help = "Define how many motifs are expected per sequence. This value is used during k-mer cutoff calculation. Default = %default meaning that there should be approximately one motif per sequence.", metavar = "double") ) opt_parser <- OptionParser(option_list = option_list, @@ -23,19 +23,19 @@ opt <- parse_args(opt_parser) #' Reduces each sequence to its most frequent region. #' #' @param input Input bed-file. Last column must be sequences. -#' @param kmer Kmer length. Default = 10 +#' @param kmer k-mer length. Default = 10 #' @param motif Estimated motif length. Default = 10 #' @param output Output file. Default = reduced.bed #' @param threads Number of threads to use. Default = 1. Use 0 for all cores. #' @param clean Delete all temporary files. -#' @param minoverlap_kmer Minimum required overlap between kmer. Used to create reduced sequence ranges out of merged kmer. Can not be greater than kmer length . Default = kmer - 1 -#' @param minoverlap_motif Minimum required overlap between motif and kmer to consider kmer significant. Used for kmer cutoff calculation. Can not be greater than motif and kmer length. Default = ceiling(motif / 2) -#' @param min_seq_length Remove sequences below this length. Defaults to the maximum value of motif and kmer and can not be lower. -#' @param motif_occurence Define how many motifs are expected per sequence. This value is used during kmer cutoff calculation. Default = 1 meaning that there should be approximately one motif per sequence. +#' @param minoverlap_kmer Minimum required overlap between k-mer. Used to create reduced sequence ranges out of merged k-mer. Can not be greater than k-mer length . Default = kmer - 1 +#' @param minoverlap_motif Minimum required overlap between motif and k-mer to consider k-mer significant. Used for k-mer cutoff calculation. Can not be greater than motif and k-mer length. Default = ceiling(motif / 2) +#' @param min_seq_length Remove sequences below this length. Defaults to the maximum value of motif and k-mer and can not be lower. +#' @param motif_occurrence Define how many motifs are expected per sequence. This value is used during k-mer cutoff calculation. Default = 1 meaning that there should be approximately one motif per sequence. #' -#' @details If there is a header supplied other then the default data.table nameing scheme ('V1', 'V2', etc.) it will be kept. +#' @details If there is a header supplied other then the default data.table naming scheme ('V1', 'V2', etc.) it will be kept. #' -reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed", threads = NULL, clean = TRUE, minoverlap_kmer = kmer - 1, minoverlap_motif = ceiling(motif / 2), min_seq_length = max(c(motif, kmer)), motif_occurence = 1) { +reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed", threads = NULL, clean = TRUE, minoverlap_kmer = kmer - 1, minoverlap_motif = ceiling(motif / 2), min_seq_length = max(c(motif, kmer)), motif_occurrence = 1) { if (system("which jellyfish", ignore.stdout = TRUE) != 0) { stop("Required program jellyfish not found! Please check whether it is installed.") } @@ -76,7 +76,7 @@ reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed" # remove sequences below minimum length if (min_seq_length < max(c(kmer, motif))) { - stop("Minimum sequence length must be greater or equal to ", max(c(motif, kmer)), " (maximum value of kmer and motif).") + stop("Minimum sequence length must be greater or equal to ", max(c(motif, kmer)), " (maximum value of k-mer and motif).") } total_rows <- nrow(bed_table) @@ -91,7 +91,7 @@ reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed" fasta_file <- paste0(basename(input), ".fasta") seqinr::write.fasta(sequences = as.list(bed_table[[ncol(bed_table)]]), names = bed_table[[4]], as.string = TRUE, file.out = fasta_file) - message("Counting kmer...") + message("Counting k-mer...") # count k-mer hashsize <- 4 ^ kmer count_output_binary <- "mer_count_binary.jf" @@ -105,20 +105,20 @@ reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed" system(command = jellyfish_dump_call, wait = TRUE) - message("Reduce kmer.") + message("Reduce k-mer.") # load mer table # columns: kmer, count kmer_counts <- data.table::fread(input = mer_count_table, header = FALSE) - # order kmer descending + # order k-mer descending data.table::setorder(kmer_counts, -V2) # compute number of hits to keep - keep_hits <- significant_kmer(bed_table, kmer = kmer, motif = motif, minoverlap = minoverlap_motif, motif_occurence = motif_occurence) + keep_hits <- significant_kmer(bed_table, kmer = kmer, motif = motif, minoverlap = minoverlap_motif, motif_occurrence = motif_occurrence) - # reduce kmer + # reduce k-mer reduced_kmer <- reduce_kmer(kmer = kmer_counts, significant = keep_hits) - message("Find kmer in sequences.") + message("Find k-mer in sequences.") # find k-mer in sequences # columns: name, start, end, width ranges_table <- find_kmer_regions(bed = bed_table, kmer_counts = reduced_kmer, minoverlap = minoverlap_kmer, threads = threads) @@ -155,21 +155,21 @@ reduce_sequence <- function(input, kmer = 10, motif = 10, output = "reduced.bed" data.table::fwrite(merged, file = output, sep = "\t", col.names = keep_col_names) } -#' Predict how many interesting kmer are possible for the given data. +#' Predict how many interesting k-mer are possible for the given data. #' #' @param bed Bed table with sequences in last column -#' @param kmer Length of kmer +#' @param kmer Length of k-mer #' @param motif Length of motif -#' @param minoverlap Minimum number of bases overlapping between kmer and motif. Must be <= motif & <= kmer. Defaults to ceiling(motif / 2). -#' @param motif_occurence Define how many motifs are expected per sequence. Default = 1 +#' @param minoverlap Minimum number of bases overlapping between k-mer and motif. Must be <= motif & <= kmer. Defaults to ceiling(motif / 2). +#' @param motif_occurrence Define how many motifs are expected per sequence. Default = 1 #' -#' @return Number of interesting kmer. -significant_kmer <- function(bed, kmer, motif, minoverlap = ceiling(motif / 2), motif_occurence = 1) { +#' @return Number of interesting k-mer. +significant_kmer <- function(bed, kmer, motif, minoverlap = ceiling(motif / 2), motif_occurrence = 1) { if (minoverlap > kmer || minoverlap > motif) { stop("Kmer & motif must be greater or equal to minoverlap!") } - if (motif_occurence <= 0) { - stop("Motif_occurence must be a numeric value above 0!") + if (motif_occurrence <= 0) { + stop("Motif_occurrence must be a numeric value above 0!") } # minimum sequence length to get all interesting overlaps @@ -180,15 +180,15 @@ significant_kmer <- function(bed, kmer, motif, minoverlap = ceiling(motif / 2), # reduce to max interesting length seq_lengths <- ifelse(seq_lengths > min_seq_length, min_seq_length, seq_lengths) - # calculate max possible kmer + # calculate max possible k-mer topx <- sum(seq_lengths - kmer + 1) - return(topx * motif_occurence) + return(topx * motif_occurrence) } -#' Orders kmer table after count descending and keeps all kmer with a cumulative sum below the given significance threshold. +#' Orders k-mer table after count descending and keeps all k-mer with a cumulative sum below the given significance threshold. #' -#' @param kmer Kmer data.table columns: kmer, count +#' @param kmer K-mer data.table columns: kmer, count #' @param significant Value from significant_kmer function. #' #' @return reduced data.table @@ -204,16 +204,16 @@ reduce_kmer <- function(kmer, significant) { #' create list of significant ranges (one for each bed entry) #' #' @param bed Data.table of bed with sequence in last column -#' @param kmer_counts Data.table of counted kmer. Column1 = kmer, column2 = count. -#' @param minoverlap Minimum overlapping nucleotides between kmers to be merged. Positive integer. Must be smaller than kmer length. +#' @param kmer_counts Data.table of counted k-mer. Column1 = kmer, column2 = count. +#' @param minoverlap Minimum overlapping nucleotides between k-mers to be merged. Positive integer. Must be smaller than k-mer length. #' @param threads Number of threads. #' #' @return Data.table with relative positions and width (start, end, width). #' -#' TODO Include number of motifs per sequence (aka motif_occurence). Attempt to keep best 2 regions for occurence = 2? Probably high impact on performance. +#' TODO Include number of motifs per sequence (aka motif_occurrence). Attempt to keep best 2 regions for occurrence = 2? Probably high impact on performance. find_kmer_regions <- function(bed, kmer_counts, minoverlap = 1 , threads = NULL) { if (nchar(kmer_counts[1, 1]) <= minoverlap) { - stop("Minoverlap must be smaller than kmer length!") + stop("Minoverlap must be smaller than k-mer length!") } names(kmer_counts)[1:2] <- c("kmer", "count") @@ -232,9 +232,9 @@ find_kmer_regions <- function(bed, kmer_counts, minoverlap = 1 , threads = NULL) return(data.table::data.table(start = NA, end = NA, width = NA, name = name)) } - # add kmer sequences + # add k-mer sequences ranges[, sub_seq := stringr::str_sub(seq, start, end)] - # add kmer count + # add k-mer count ranges[, count := kmer_counts[ranges[["sub_seq"]], "count", on = "kmer"]] #### reduce ranges @@ -253,7 +253,7 @@ find_kmer_regions <- function(bed, kmer_counts, minoverlap = 1 , threads = NULL) which(member == x) }) - # calculate component score (= sum of kmer count) + # calculate component score (= sum of k-mer count) score <- vapply(node_membership, FUN.VALUE = numeric(1), function(x) { sum(kmer_counts[x, "count"]) }) From d86f788766e24a2ee3c5144cd2273d8a491b9de2 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Thu, 20 Dec 2018 13:57:34 +0100 Subject: [PATCH 25/33] fixed more typos --- bin/reduce_sequence.R | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/bin/reduce_sequence.R b/bin/reduce_sequence.R index cf91187..53ac69f 100644 --- a/bin/reduce_sequence.R +++ b/bin/reduce_sequence.R @@ -3,7 +3,7 @@ library("optparse") option_list <- list( make_option(opt_str = c("-i", "--input"), default = NULL, help = "Input bed-file. Last column must be sequences.", metavar = "character"), - make_option(opt_str = c("-k", "--kmer"), default = 10, help = "k-mer length. Default = %default", metavar = "integer"), + make_option(opt_str = c("-k", "--kmer"), default = 10, help = "K-mer length. Default = %default", metavar = "integer"), make_option(opt_str = c("-m", "--motif"), default = 10, help = "Estimated motif length. Default = %default", metavar = "integer"), make_option(opt_str = c("-o", "--output"), default = "reduced.bed", help = "Output file. Default = %default", metavar = "character"), make_option(opt_str = c("-t", "--threads"), default = 1, help = "Number of threads to use. Use 0 for all available cores. Default = %default", metavar = "integer"), @@ -28,7 +28,7 @@ opt <- parse_args(opt_parser) #' @param output Output file. Default = reduced.bed #' @param threads Number of threads to use. Default = 1. Use 0 for all cores. #' @param clean Delete all temporary files. -#' @param minoverlap_kmer Minimum required overlap between k-mer. Used to create reduced sequence ranges out of merged k-mer. Can not be greater than k-mer length . Default = kmer - 1 +#' @param minoverlap_kmer Minimum required overlap between k-mer. Used to create reduced sequence ranges out of merged k-mer. Can not be greater than k-mer length. Default = kmer - 1 #' @param minoverlap_motif Minimum required overlap between motif and k-mer to consider k-mer significant. Used for k-mer cutoff calculation. Can not be greater than motif and k-mer length. Default = ceiling(motif / 2) #' @param min_seq_length Remove sequences below this length. Defaults to the maximum value of motif and k-mer and can not be lower. #' @param motif_occurrence Define how many motifs are expected per sequence. This value is used during k-mer cutoff calculation. Default = 1 meaning that there should be approximately one motif per sequence. From 756e98f550ac6dde7df603fb506a83d203654182 Mon Sep 17 00:00:00 2001 From: Schultheis Date: Thu, 20 Dec 2018 14:01:22 +0100 Subject: [PATCH 26/33] process description for reduce_sequence and clustering --- pipeline.nf | 2 ++ 1 file changed, 2 insertions(+) diff --git a/pipeline.nf b/pipeline.nf index db3bbdf..c5f16b6 100644 --- a/pipeline.nf +++ b/pipeline.nf @@ -267,6 +267,7 @@ process overlap_with_known_TFBS { /* +Reduce each sequence to its most conserved region. */ process reduce_sequence { conda "${path_env}" @@ -287,6 +288,7 @@ process reduce_sequence { /* +Cluster all sequences. Appends a column with cluster numbers to the bed-file. */ process clustering { conda "${path_env}" From 5e462666c3e84c1173443d5b4f93470ee6adbb19 Mon Sep 17 00:00:00 2001 From: renewiegandt Date: Thu, 20 Dec 2018 14:11:14 +0100 Subject: [PATCH 27/33] Fixed typo in bed_to_fasta.R --- bin/bed_to_fasta.R | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/bin/bed_to_fasta.R b/bin/bed_to_fasta.R index 73ea732..e0ade14 100644 --- a/bin/bed_to_fasta.R +++ b/bin/bed_to_fasta.R @@ -13,7 +13,7 @@ opt_parser <- OptionParser(option_list = option_list, opt <- parse_args(opt_parser) #' Splitting BED-files depending on their cluster. -#' The Sequences of each cluster are writen as an FASTA-file. +#' The Sequences of each cluster are written as an FASTA-file. #' @param bedInput BED-file with sequences and cluster-id as last two columns: #' Sequence: second last column; Cluster ID: last column #' @param prefix prefix for filenames From 1c6bcf120bdd0832d6df3061584031cf4b3a16b2 Mon Sep 17 00:00:00 2001 From: renewiegandt Date: Thu, 20 Dec 2018 14:52:37 +0100 Subject: [PATCH 28/33] Added new parameter list to README.mf --- README.md | 16 ++++++++-------- 1 file changed, 8 insertions(+), 8 deletions(-) diff --git a/README.md b/README.md index 41ee001..d139f97 100644 --- a/README.md +++ b/README.md @@ -11,7 +11,7 @@ For further information read the [documentation](https://github.molgen.mpg.de/lo ## Installation Start with installing all dependencies listed above (Nextflow, conda) and downloading all files from the [GitHub repository](https://github.molgen.mpg.de/loosolab/masterJLU2018). -Every other dependency will be automatically installed by Nextflow using conda. For that a new conda enviroment will be created, which can be found in the from Nextflow created work directory after the first pipeline run. +Every other dependency will be automatically installed by Nextflow using conda. For that a new conda enviroment will be created, which can be found in the from Nextflow created work directory after the first pipeline run. It is **not** required to create and activate the enviroment from the yaml-file beforehand. **Important Note:** For conda the channel bioconda needs to be set as highest priority! This is required due to two differnt packages with the same name in different channels. For the pipeline the package jellyfish from the channel bioconda is needed and **NOT** the jellyfisch package from the channel conda-forge! @@ -30,14 +30,16 @@ Required arguments: --genome_fasta Path to genome in FASTA-format --motif_db Path to motif-database in MEME-format --config Path to UROPA configuration file - --create_known_tfbs_path Path to directory where output from tfbsscan (known motifs) are stored. - Path can be set as tfbs_path in next run. (Default: './') - --out Output Directory (Default: './out/') - + --organism Input organism [hg38 | hg19 | mm9 | mm10] + --out Output Directory (Default: './out/') + Optional arguments: - + --help [0|1] 1 to show this help message. (Default: 0) --tfbs_path Path to directory with output from tfbsscan. If given tfbsscan will not be run. + --create_known_tfbs_path Path to directory where output from tfbsscan (known motifs) are stored. + Path can be set as tfbs_path in next run. (Default: './') + --gtf_path Path to gtf-file. If path is set the process which creats a gtf-file is skipped. Footprint extraction: --window_length INT This parameter sets the length of a sliding window. (Default: 200) @@ -77,7 +79,6 @@ Optional arguments: --motif_similarity_thresh FLOAT Threshold for motif similarity score (Default: 0.00001) Creating GTF: - --organism [hg38 | hg19 | mm9 | mm10] Input organism --tissues List/String List of one or more keywords for tissue-/category-activity, categories must be specified as in JSON config All arguments can be set in the configuration files @@ -103,4 +104,3 @@ path=[Path to given masterenv.yml file] conda env create --name masterenv -f $path ``` When the enviroment is created, set the variable 'path_env' in the configuration file as the path to it. - From 13bccdafbb5ab5d4c8f2e43304167ad564a50106 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ren=C3=A9=20Wiegandt?= Date: Fri, 21 Dec 2018 06:46:11 -0500 Subject: [PATCH 29/33] Fixed bug in pipeline.nf: parameter gtf_path is now working --- pipeline.nf | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/pipeline.nf b/pipeline.nf index 1be18be..f5f709c 100644 --- a/pipeline.nf +++ b/pipeline.nf @@ -9,7 +9,7 @@ params.tfbs_path="" params.create_known_tfbs_path = "./" params.help = 0 - params.get_path="" + params.gtf_path="" params.out = "./out/" //peak_calling @@ -583,7 +583,7 @@ process create_GTF { file ('*.gtf') into gtf when: - gtf_path == "" + params.gtf_path == "" script: """ @@ -591,7 +591,7 @@ process create_GTF { """ } -if (gtf_path == "") { +if (params.gtf_path == "") { gtf_for_uropa = gtf } else { gtf_for_uropa = Channel.fromPath(params.gtf_path) From b7c80c85596c25cdfa10c69f315242d4ebbc07c6 Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Ren=C3=A9=20Wiegandt?= Date: Thu, 3 Jan 2019 07:48:52 -0500 Subject: [PATCH 30/33] sorting scripts depending on their function --- .../footprints_extraction.py | 0 bin/{ => 1.2_filter_motifs}/compareBed.sh | 0 bin/{ => 1.2_filter_motifs}/maxScore.R | 0 bin/{ => 1.2_filter_motifs}/merge.R | 0 bin/{ => 1.2_filter_motifs}/tfbsscan.py | 0 bin/{ => 2.1_clustering}/cdhit_wrapper.R | 0 bin/{ => 2.1_clustering}/reduce_sequence.R 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