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author | oinizan |
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date | Wed, 18 Oct 2017 05:30:40 -0400 |
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#!/usr/bin/env python2.7 # # Copyright (C) 2014 INRA # # This program is free software: you can redistribute it and/or modify # it under the terms of the GNU General Public License as published by # the Free Software Foundation, either version 3 of the License, or # (at your option) any later version. # # This program is distributed in the hope that it will be useful, # but WITHOUT ANY WARRANTY; without even the implied warranty of # MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the # GNU General Public License for more details. # # You should have received a copy of the GNU General Public License # along with this program. If not, see <http://www.gnu.org/licenses/>. # __author__ = 'Plateforme bioinformatique Toulouse and SIGENAE' __copyright__ = 'Copyright (C) 2015 INRA' __license__ = 'GNU General Public License' __version__ = '1.2.0' __email__ = 'frogs@inra.fr' __status__ = 'prod' import os import sys import gzip import time import tarfile import argparse CURRENT_DIR = os.path.dirname(os.path.abspath(__file__)) # PATH BIN_DIR = os.path.abspath(os.path.join(os.path.dirname(CURRENT_DIR), "libexec")) os.environ['PATH'] = BIN_DIR + os.pathsep + os.environ['PATH'] # PYTHONPATH LIB_DIR = os.path.abspath(os.path.join(os.path.dirname(CURRENT_DIR), "lib")) sys.path.append(LIB_DIR) if os.getenv('PYTHONPATH') is None: os.environ['PYTHONPATH'] = LIB_DIR else: os.environ['PYTHONPATH'] = os.environ['PYTHONPATH'] + os.pathsep + LIB_DIR from frogsUtils import * ################################################################################################################################################## # # COMMAND LINES # ################################################################################################################################################## class Demultiplex(Cmd): """ @summary : Demultiplex samples. """ def __init__(self, R1_input_file, R2_input_file, barcode_file, mismatches, end, global_tmp_files, R1_output_files, R2_output_files, demultiplex_err_files1, demultiplex_err_files2, demultiplex_log): """ @param R1_input_file : [str] Path to the R1 fastq file. @param R2_input_file : [str] Path to the R2 fastq file. @param barcode_file : [str] Path to barcodes and samples (one line by sample) description file. Line format : SAMPLE_NAME<TAB>BARCODE. @param mismatches : [int] Number of mismatches allowed @param end : [str] barcode ends ? forward : bol or reverse : eol (def bol) @param global_tmp_files : [str] Path for R1 and R2 files. @param R1_output_files : [list] Paths to the R1 fastq files (one by sample). User provides an empty list. @param R2_output_files : [list] Paths to the R2 fastq files (one by sample). User provides an empty list. @param demultiplex_err_files : [list] Paths to the files with ambiguous and unmatched reads. User provides an empty list. """ tmp_files = TmpFiles( global_tmp_files.tmp_dir ) tmp_folder = os.path.join( global_tmp_files.tmp_dir, global_tmp_files.prefix + "_tmp", tmp_files.prefix ) global_tmp_files.dirs.append(tmp_folder) if not os.path.exists(tmp_folder): os.mkdir(tmp_folder) self.samples_names = list() # Update output data FH_barcode = open( barcode_file ) for line in FH_barcode: sample_name, barcode = line.strip().rsplit(None, 1) R1_output_files.append( os.path.join(tmp_folder, sample_name + '_R1.fastq') ) global_tmp_files.files.append(os.path.join(tmp_folder, sample_name + '_R1.fastq') ) if R2_input_file != None: R2_output_files.append( os.path.join(tmp_folder, sample_name + '_R2.fastq') ) global_tmp_files.files.append(os.path.join(tmp_folder, sample_name + '_R2.fastq')) self.samples_names.append( sample_name.replace(' ', '_') ) FH_barcode.close() self.R1_input_file = R1_input_file self.ambiguous = os.path.join(tmp_folder, 'ambiguous_R1.fastq') self.unmatched = os.path.join(tmp_folder, 'unmatched_R1.fastq') demultiplex_err_files1.extend( [self.ambiguous,self.unmatched] ) global_tmp_files.files.extend( [self.ambiguous,self.unmatched] ) if R2_input_file != None: demultiplex_err_files2.extend( [os.path.join(tmp_folder, 'ambiguous_R2.fastq'),os.path.join(tmp_folder, 'unmatched_R2.fastq') ]) global_tmp_files.files.extend( [os.path.join(tmp_folder, 'ambiguous_R2.fastq'),os.path.join(tmp_folder, 'unmatched_R2.fastq') ]) # Set class if R2_input_file != None: Cmd.__init__( self, 'perl ' + os.environ['TOOL_DIREDCTORY'] + '/splitbc.pl', 'Demultiplex reads.', R1_input_file + ' ' + R2_input_file + ' --' + end + ' --bcfile ' + barcode_file + ' --mismatches ' + `mismatches` + ' --trim --no_adapt --prefix-r1 ' + os.path.join(tmp_folder, '%_R1.fastq') +\ ' --prefix-r2 ' + os.path.join(tmp_folder, '%_R2.fastq') + ' >> ' + demultiplex_log, None ) else: Cmd.__init__( self, 'perl ' + os.environ['TOOL_DIRECTORY'] + '/splitbc.pl', 'Demultiplex reads.', R1_input_file + ' --' + end + ' --bcfile ' + barcode_file + ' --mismatches ' + `mismatches` + ' --trim --no_adapt --prefix-r1 ' + os.path.join(tmp_folder, '%_R1.fastq') +\ ' >> ' + demultiplex_log, None ) def parser(self, log_file): """ @summary : Parse the command results to add information in log_file. @log_file : [str] Path to the sample process log file. """ # Parse output nb_seq_before = get_fastq_nb_seq(self.R1_input_file) nb_seq_unmatched = get_fastq_nb_seq(self.unmatched) nb_seq_ambiguous = get_fastq_nb_seq(self.ambiguous) # Write result FH_log = Logger( log_file ) FH_log.write( 'Results :\n' ) FH_log.write( '\tnb seq before demultiplexing : ' + str(nb_seq_before) + '\n' ) FH_log.write( '\tnb seq after process matched : ' + str(nb_seq_before - nb_seq_unmatched) + '\n' ) FH_log.write( '\tnb seq after process non-ambiguous : ' + str(nb_seq_before - nb_seq_unmatched - nb_seq_ambiguous) + '\n' ) FH_log.close() def get_version(self): """ @summary : Returns the program version number. @return : version number if this is possible, otherwise this method return 'unknown'. """ return Cmd.get_version(self, 'stdout') class Archive(Cmd): """ @summary : Creates an archive with files. """ def __init__(self, archived_files, archive_path): """ @param archived_files: [list] Files added in final archive. @param archive_path: [str] Path to the new archive. """ tmp_files=TmpFiles( os.path.dirname(archive_path) ) tmp_folder = os.path.join( tmp_files.tmp_dir, tmp_files.prefix) tmp_files.dirs.append(tmp_folder) if not os.path.exists(tmp_folder): os.makedirs(tmp_folder) if len(archived_files) == 0: raise Exception( "At least one file must be add to the archive '" + archive_path + "'." ) archived_basenames = list() for current in archived_files: if not os.path.dirname(current) == tmp_folder: os.rename(current, os.path.join(tmp_folder,os.path.basename(current))) tmp_files.files.append(os.path.join(tmp_folder,os.path.basename(current))) archived_basenames.append(os.path.basename(current)) Cmd.__init__( self, 'tar', 'Archives files.', '-zcf ' + archive_path + ' -C ' + tmp_folder + " " + " ".join(archived_basenames), None ) self.Files=tmp_files def parser(self,log_file): self.Files.deleteAll() ################################################################################################################################################## # # FUNCTIONS # ################################################################################################################################################## def is_gzip( file ): """ @return: [bool] True if the file is gziped. @param file : [str] Path to processed file. """ is_gzip = None FH_input = gzip.open( file ) try: FH_input.readline() is_gzip = True except: is_gzip = False finally: FH_input.close() return is_gzip def split_barcode_file( barcode_file, barcodes_file_list, global_tmp_files ): """ @summary: In case of double multiplexe, split barcode file in one forward and multiple reverse barcode files @param barcode_file: [str] Path to the input barcode file @param barcodes_file_list: [list] List of path to the ouput barcode files @param out_dir: [str] path to the output directory to write barcode files """ out_dir = global_tmp_files.tmp_dir barcode_input = open(barcode_file,"r") barcode_dict={} for l in barcode_input.readlines(): [s,f,r]=l.strip().split() if not "forward_bc" in barcode_dict: barcode_dict["forward_bc"] = [f+"\t"+f] elif not f+"\t"+f in barcode_dict["forward_bc"]: barcode_dict["forward_bc"].append( f+"\t"+f) if not f+"_reverse_bc" in barcode_dict: barcode_dict[f+"_reverse_bc"] = [s+"\t"+r] else : barcode_dict[f+"_reverse_bc"].append(s+"\t"+r) f=barcode_dict.pop("forward_bc") barcodes_file_list.append(os.path.join(out_dir,"forward_bc")) global_tmp_files.files.append(os.path.join(out_dir,"forward_bc")) FH_out = open(os.path.join(out_dir,"forward_bc"),"w") FH_out.write("\n".join(f)+"\n") FH_out.close() for bc_file in barcode_dict: barcodes_file_list.append(os.path.join(out_dir,bc_file)) global_tmp_files.files.append(os.path.join(out_dir,bc_file)) FH_out = open(os.path.join(out_dir,bc_file),"w") FH_out.write("\n".join(barcode_dict[bc_file])+"\n") FH_out.close() def get_fastq_nb_seq( fastq_file ): """ @summary: Returns the number of sequences in fastq_file. @param fastq_file: [str] Path to the fastq file processed. @return: [int] The number of sequences. """ FH_input = None if not is_gzip(fastq_file): FH_input = open( fastq_file ) else: FH_input = gzip.open( fastq_file ) nb_line = 0 for line in FH_input: nb_line += 1 FH_input.close() nb_seq = nb_line/4 return nb_seq def concat_files(list_input, output_file): FH_out=open(output_file,"w") for f in list_input : FH_in = open(f) string="" i=0 for line in FH_in: string+= line i+=1 if i==2000 : FH_out.write(string) string="" i=0 if i != 0: FH_out.write(string) FH_in.close() FH_out.close() def summarise_results( summary_file, barcode_file, log_file ): """ @summary: Writes one summary of results from several logs. @param summary_file: [str] The output file. @param log_files: [list] The list of path to log files (one log file by sample). """ sample_dict=dict() FH_barcode= open(barcode_file) for line in FH_barcode: sample_dict[line.split()[0]]=0 FH_summary = open(summary_file, "w") FH_summary.write( "#sample\tcount\n") FH_log = open(log_file,"r") sample_dict["unmatched"]=0 sample_dict["ambiguous"]=0 for line in FH_log.readlines(): if line.startswith("Barcode") or line.startswith("total") : pass else : l=line.replace('(','\t').split() if l[0] in sample_dict: sample_dict[l[0]] += int(l[1]) for s in sample_dict: FH_summary.write(s + '\t' + str(sample_dict[s]) + '\n') FH_summary.close() ################################################################################################################################################## # # MAIN # ################################################################################################################################################## if __name__ == "__main__": # Manage parameters parser = argparse.ArgumentParser( description='Split by samples the reads in function of inner barcode.' ) parser.add_argument('-m', '--mismatches', type=int, default=0, help="Number of mismatches allowed in barcode. [Default: %(default)s]") parser.add_argument('-e', '--end', type=str, default="bol", help="barcode is at the begining of the forward end (bol) or of the reverse (eol) or both (both). [Default: %(default)s]") parser.add_argument( '--debug', default=False, action='store_true', help="Keep temporary files to debug program." ) parser.add_argument( '-v', '--version', action='version', version=__version__ ) # Inputs group_input = parser.add_argument_group( 'Inputs' ) group_input.add_argument( '--input-R1', required=True, help='The R1 sequence file with all samples (format: fastq).' ) group_input.add_argument( '--input-R2', default=None, help='The R2 sequence file with all samples (format: fastq).' ) group_input.add_argument( '--input-barcode', help='This file describes barcodes and samples (one line by sample). Line format : SAMPLE_NAME<TAB>BARCODE or SAMPLE_NAME<TAB>BARCODE_FW<TAB>BARCODE_RV.' ) group_output = parser.add_argument_group( 'Outputs' ) # Outputs group_output.add_argument( '--output-demultiplexed', default="demultiplexed_read.tar.gz", help='The tar file containing R1 files and R2 files for each sample (format: tar). [Default: %(default)s]' ) group_output.add_argument( '--output-excluded', default="undemultiplexed_read.tar.gz", help='The tar file containing R1 files and R2 files not demultiplexed (format: tar). [Default: %(default)s]' ) group_output.add_argument( '-s', '--summary', default='summary.tsv', help='TSV file with summary of filters results (format: TSV). [Default: %(default)s]') group_output.add_argument( '-l', '--log-file', default=sys.stdout, help='This output file will contain several information on executed commands.') args = parser.parse_args() prevent_shell_injections(args) Logger.static_write(args.log_file, "## Application\nSoftware :" + sys.argv[0] + " (version : " + str(__version__) + ")\nCommand : " + " ".join(sys.argv) + "\n\n") # Process R1_files = list() R2_files = list() tmp_barcode_files = list() tmp_R1_files = list() tmp_R2_files = list() demultiplex_err_files1 = list() demultiplex_err_files2 = list() excluded_R1_file = os.path.join(os.path.split(args.output_demultiplexed)[0],os.path.basename(args.input_R1)+"_excluded_demult") if args.input_R2 != None : excluded_R2_file = os.path.join(os.path.split(args.output_demultiplexed)[0],os.path.basename(args.input_R2)+"_excluded_demult") uniq_id = str(time.time()) + "_" + str(os.getpid()) tmp_files = TmpFiles( os.path.split(args.output_demultiplexed)[0] ) demultiplex_log = tmp_files.add("Demult.log") tmp_folder=tmp_files.add_dir("tmp") os.mkdir(tmp_folder) sample_list=[] try: # Process if args.end == "bol" or args.end == "eol" : info="\n#Demultiplexing " + os.path.basename(args.input_R1) if args.input_R2 != None: info+= " and " + os.path.basename(args.input_R2) info += " with " + os.path.basename(args.input_barcode) + " in " + args.end + " strand\n" Logger.static_write(args.log_file,info) Demultiplex(args.input_R1, args.input_R2, args.input_barcode, args.mismatches, args.end, tmp_files, R1_files, R2_files, demultiplex_err_files1,demultiplex_err_files2, demultiplex_log).submit( args.log_file ) else: split_barcode_file(args.input_barcode, tmp_barcode_files, tmp_files) info="\n#Demultiplexing " + os.path.basename(args.input_R1) if args.input_R2 != None: info+= " and " + os.path.basename(args.input_R2) info += " with " + os.path.basename(tmp_barcode_files[0]) + " in bol strand\n" Logger.static_write(args.log_file,info) Demultiplex(args.input_R1, args.input_R2, tmp_barcode_files[0], args.mismatches, "bol", tmp_files, tmp_R1_files, tmp_R2_files, demultiplex_err_files1,demultiplex_err_files2, demultiplex_log).submit( args.log_file ) for idx,read1_file in enumerate(tmp_R1_files): bc = os.path.basename(read1_file).replace("_R1.fastq","") if os.path.join(tmp_files.tmp_dir,bc+"_reverse_bc") in tmp_barcode_files: if os.stat(tmp_R1_files[idx]).st_size != 0 : info="\n#Demultiplexing " + os.path.basename(tmp_R1_files[idx]) if args.input_R2 != None: info+= " and " + os.path.basename(tmp_R2_files[idx]) info += " with " + bc+"_reverse_bc" + " in eol strand\n" Logger.static_write(args.log_file,info) if args.input_R2 != None: Demultiplex(tmp_R1_files[idx], tmp_R2_files[idx], os.path.join(tmp_files.tmp_dir,bc+"_reverse_bc"), args.mismatches, "eol", tmp_files, R1_files, R2_files, demultiplex_err_files1, demultiplex_err_files2, demultiplex_log).submit( args.log_file ) else: Demultiplex(tmp_R1_files[idx], None, os.path.join(tmp_files.tmp_dir,bc+"_reverse_bc"), args.mismatches, "eol", tmp_files, R1_files, R2_files, demultiplex_err_files1, demultiplex_err_files2, demultiplex_log).submit( args.log_file ) Logger.static_write(args.log_file,"\n#Summarising result\n") summarise_results( args.summary, args.input_barcode, demultiplex_log ) Logger.static_write(args.log_file,"\n#Concatenation of undemultiplexed files 1\n") concat_files(demultiplex_err_files1, excluded_R1_file ) if len(R2_files) > 0: Logger.static_write(args.log_file,"\n#Concatenation of undemultiplexed files 2\n") concat_files(demultiplex_err_files2, excluded_R2_file ) Logger.static_write(args.log_file,"\n#Archive demultiplexed R1 and R2 files\n") Archive(R1_files + R2_files, args.output_demultiplexed).submit( args.log_file ) Logger.static_write(args.log_file,"\n#Archive undemultiplexed R1 and R2 files\n") Archive([excluded_R1_file,excluded_R2_file], args.output_excluded).submit( args.log_file ) else: Logger.static_write(args.log_file,"\n#Archive demultiplexed files\n") Archive(R1_files, args.output_demultiplexed).submit( args.log_file ) Logger.static_write(args.log_file,"\n#Archive undemultiplexed files\n") Archive([excluded_R1_file], args.output_excluded).submit( args.log_file ) # Remove temporary files finally: if not args.debug: Logger.static_write(args.log_file,"\n#Removing temporary files\n") tmp_files.deleteAll()