This repository has been archived by the owner on Jun 26, 2023. It is now read-only.
-
Notifications
You must be signed in to change notification settings - Fork 1
/
ptr.py
executable file
·408 lines (348 loc) · 22.8 KB
/
ptr.py
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
83
84
85
86
87
88
89
90
91
92
93
94
95
96
97
98
99
100
101
102
103
104
105
106
107
108
109
110
111
112
113
114
115
116
117
118
119
120
121
122
123
124
125
126
127
128
129
130
131
132
133
134
135
136
137
138
139
140
141
142
143
144
145
146
147
148
149
150
151
152
153
154
155
156
157
158
159
160
161
162
163
164
165
166
167
168
169
170
171
172
173
174
175
176
177
178
179
180
181
182
183
184
185
186
187
188
189
190
191
192
193
194
195
196
197
198
199
200
201
202
203
204
205
206
207
208
209
210
211
212
213
214
215
216
217
218
219
220
221
222
223
224
225
226
227
228
229
230
231
232
233
234
235
236
237
238
239
240
241
242
243
244
245
246
247
248
249
250
251
252
253
254
255
256
257
258
259
260
261
262
263
264
265
266
267
268
269
270
271
272
273
274
275
276
277
278
279
280
281
282
283
284
285
286
287
288
289
290
291
292
293
294
295
296
297
298
299
300
301
302
303
304
305
306
307
308
309
310
311
312
313
314
315
316
317
318
319
320
321
322
323
324
325
326
327
328
329
330
331
332
333
334
335
336
337
338
339
340
341
342
343
344
345
346
347
348
349
350
351
352
353
354
355
356
357
358
359
360
361
362
363
364
365
366
367
368
369
370
371
372
373
374
375
376
377
378
379
380
381
382
383
384
385
386
387
388
389
390
391
392
393
394
395
396
397
398
399
400
401
402
403
404
405
406
407
408
__author__ = 'alipirani'
import sys
import os
import argparse
import errno
from datetime import datetime
import ConfigParser
from config_settings import ConfigSectionMap
from modules.check_subroutines import *
from modules.stages import *
from modules.bedtools import *
from modules.qualimap import *
if sys.version_info < (3, 2):
import subprocess32 as sp
else:
import subprocess as sp
from modules.logging_subprocess import *
from modules.log_modules import *
from modules.generate_PTR_dataframe import *
from argparse import RawTextHelpFormatter
# Command Line Argument Parsing
def parser():
parser = argparse.ArgumentParser(description='GROWTH RATE ANALYSIS PIPELINE. \n\nThis pipeline calculates peak to through ratio(PTR; ratio of copy numbers at origin of replication to terminus) using sequencing read coverage.', formatter_class=RawTextHelpFormatter)
required = parser.add_argument_group('Required arguments')
optional = parser.add_argument_group('Optional arguments')
required.add_argument('-type', action='store', dest="type", help='Type of Sequencing Reads: SE or PE', required=True)
optional.add_argument('-config', action='store', dest="config", help='Path to Config file', required=False)
required.add_argument('-PE1', action='store', dest="forward_raw", help='Path to paired-end file 1', required=True)
optional.add_argument('-PE2', action='store', dest="reverse_raw", help='Path to paired-end file 2', required=False)
required.add_argument('-steps', action='store', dest="steps", help='Growth-Rate-Analysis Steps in sequential order.\n\n'
'1. "All": This option will run all the steps starting from: \n\n - cleaning reads using Trimmomatic, \n - Aligning reads using Bowtie, \n - Post-align modifications using samtools, \n - Bedtools to extract coverage across the genome and \n - PTR calculation from Bed coverage;\n\n'
'2. "clean,align,post-align,coverage,ptr" : This option will also run all steps starting from: \n\n - cleaning reads using Trimmomatic, \n - Aligning reads using Bowtie, \n - Post-align modifications using samtools, \n - Bedtools to extract coverage across the genome and \n - PTR calculation from Bed coverage;\n\nIMPORTANT: The order is required to be sequential while using this option. \nAlso, while skipping any of the step, make sure you have results for previous steps already present in your output folder.\n\n')
required.add_argument('-o', action='store', dest="output_folder", help='Output Path ending with output directory name to save the results', required=True)
required.add_argument('-analysis', action='store', dest="analysis_name", help='Unique analysis name to save the results', required=True)
required.add_argument('-index', action='store', dest="index", help='Reference Index Name. Change this argument in config file and mention the reference header name such as KP_NTUH_chr/KPNIH1/KPNIH32.', required=True)
optional.add_argument('-adapter', action='store', dest="adapter", help='Path to adapter fasta file')
optional.add_argument('-c', action='store', dest="croplength", help='Crop Length in case needed')
#optional.add_argument('-f', action='store', dest="bam_input", help='Input Bam')
return parser
# Main Pipeline
def pipeline(args, logger):
keep_logging('\nSTART: Pipeline\n', 'START: Pipeline\n', logger, 'info')
""" SANITATION CHECKS """
# Check Subroutines for Arguments, Input FASTQ files, Reference Index
keep_logging('START: Checking Dependencies...', 'Checking Dependencies', logger, 'info')
# Reference Genome file name
reference = ConfigSectionMap(args.index, Config)['ref_path'] + "/" + ConfigSectionMap(args.index, Config)['ref_name']
keep_logging('Getting Reference Genome name from config file: {}'.format(reference), 'Getting Reference Genome name from config file: {}'.format(reference), logger, 'info')
# # Check IF FASTQ files exists
# if args.type != "PE" and args.type != "BAM":
# reverse_raw = "None"
# file_exists(args.forward_raw, args.forward_raw, reference)
# elif args.type != "PE" and args.type != "SE":
# print "BAM type... Not Integrated... continue"
# else:
# file_exists(args.forward_raw, args.reverse_raw, reference)
# Check Java Version
java_check()
keep_logging('END: Checking Dependencies...\n', 'END: Checking Dependencies\n', logger, 'info')
""" INDIVIDUAL SUBPROCESS FOR EACH PIPELINE STEPS"""
## 1. Pre-Processing Raw reads using Trimmomatic
def clean():
keep_logging('START: Pre-Processing Raw reads using Trimmomatic',
'START: Pre-Processing Raw reads using Trimmomatic', logger, 'info')
if args.type == "PE":
# print "skip trimming"
trimmomatic(args.forward_raw, args.reverse_raw, args.output_folder, args.croplength, args.adapter, logger, Config)
else:
reverse_raw = "None"
# print "skip trimming"
trimmomatic(args.forward_raw, reverse_raw, args.output_folder, args.croplength, args.adapter, logger, Config)
keep_logging('END: Pre-Processing Raw reads using Trimmomatic\n',
'END: Pre-Processing Raw reads using Trimmomatic\n', logger, 'info')
## 2. Stages: prepare readgroups and map reads to a reference genome using user-provided aligner as set in config file.
def align_reads():
keep_logging('START: Mapping Reads using BWA', 'START: Mapping Reads using BWA', logger, 'info')
split_field = prepare_readgroup(args.forward_raw, ConfigSectionMap("pipeline", Config)['aligner'], logger)
out_sam = align(args.output_folder, args.index, split_field, args.analysis_name, files_to_delete, logger, Config, args.type)
keep_logging('END: Mapping Reads using BWA', 'END: Mapping Reads using BWA', logger, 'info')
return out_sam
## 3. Stages: Post-Alignment using SAMTOOLS, PICARD etc
def post_align(out_sam):
keep_logging('START: Post-Alignment using SAMTOOLS, PICARD etc...',
'START: Post-Alignment using SAMTOOLS, PICARD etc...', logger, 'info')
out_sorted_bam = prepare_bam(out_sam, args.output_folder, args.analysis_name, files_to_delete, logger, Config)
keep_logging('END: Post-Alignment using SAMTOOLS, PICARD etc...',
'END: Post-Alignment using SAMTOOLS, PICARD etc...', logger, 'info')
# out_sorted_bam = "%s/%s_aln_sort.bam" % (args.output_folder, args.analysis_name)
return out_sorted_bam
## 4. Stages: Generate Bedgrapg Coverage File for PTR calculation
def bedgraph(out_sorted_bam):
final_coverage_file = bedtools(out_sorted_bam, args.output_folder, args.analysis_name, logger, Config)
#final_coverage_file = "%s/%s_coverage.bed" % (args.output_folder, args.analysis_name)
return final_coverage_file
## 5. Stages: PTR Analysis
def ptr(final_coverage_file):
keep_logging('START: Analyzing Bedfiles for PTR analysis', 'START: Analyzing Bedfiles for PTR analysis', logger,
'info')
generate_PTR_dataframe(final_coverage_file, args.output_folder, args.analysis_name, logger, Config)
keep_logging('END: Analyzing Bedfiles for PTR analysis\n', 'END: Analyzing Bedfiles for PTR analysis\n', logger,
'info')
## 6. Stages: Statistics
def stats(out_sorted_bam):
keep_logging('START: Generating Statistics Reports', 'START: Generating Statistics Reports', logger, 'info')
alignment_stats_file = alignment_stats(out_sorted_bam, args.output_folder, args.analysis_name, logger, Config)
<<<<<<< HEAD:ptr.py
=======
# Commented out. These methods are for debugging and troubleshooting purpose
>>>>>>> 82a9fde8f5c26edb4007075eb056584348a71e72:pipeline.py
# gatk_DepthOfCoverage(out_sorted_bam, args.output_folder, args.analysis_name, reference, logger, Config)
# final_coverage_file = "%s/%s_coverage.bed" % (args.output_folder, args.analysis_name)
# keep_logging('END: Generating Statistics Reports\n', 'END: Generating Statistics Reports\n', logger, 'info')
# final_coverage_file = "%s/%s_coverage.bed" % (args.output_folder, args.analysis_name)
""" Start the pipeline: """
# Split comma-seperated values provided with -steps argument and decide the starting point of pipeline.
steps_list = args.steps.split(',')
# Decide the starting point of pipeline and run accordingly.
if len(steps_list) == 1:
if steps_list[0] == "All":
clean()
out_sam = align_reads()
out_sorted_bam = post_align(out_sam)
<<<<<<< HEAD:ptr.py
final_coverage_file = bedgraph(out_sorted_bam)
stats(out_sorted_bam)
ptr(final_coverage_file)
=======
final_coverage_file = bedgraph(out_sorted_bam)
stats(out_sorted_bam)
final_coverage_file = bedgraph(out_sorted_bam)
stats(out_sorted_bam)
ptr(final_coverage_file)
>>>>>>> 82a9fde8f5c26edb4007075eb056584348a71e72:pipeline.py
if steps_list[0] == "ptr":
final_coverage_file = "%s/%s_coverage.bed" % (args.output_folder, args.analysis_name)
out_sorted_bam = "%s/%s_aln_sort.bam" % (args.output_folder, args.analysis_name)
final_coverage_file = "%s/%s_coverage.bed" % (args.output_folder, args.analysis_name)
stats(out_sorted_bam)
ptr(final_coverage_file)
# Run individual pipeline steps based on the first value found in steps_list array: clean, align, post-align, bedgraph, ptr, stats etc
else:
if steps_list[0] == "clean":
clean()
out_sam = align_reads()
out_sorted_bam = post_align(out_sam)
final_coverage_file = bedgraph(out_sorted_bam)
stats(out_sorted_bam)
ptr(final_coverage_file)
elif steps_list[0] == "align":
out_sam = align_reads()
out_sorted_bam = post_align(out_sam)
final_coverage_file = bedgraph(out_sorted_bam)
stats(out_sorted_bam)
ptr(final_coverage_file)
elif steps_list[0] == "post-align":
out_sam = "%s/%s_aln.sam" % (args.output_folder, args.analysis_name)
out_sorted_bam = post_align(out_sam)
final_coverage_file = bedgraph(out_sorted_bam)
stats(out_sorted_bam)
ptr(final_coverage_file)
elif steps_list[0] == "bedgraph":
out_sorted_bam = "%s/%s_aln_sort.bam" % (args.output_folder, args.analysis_name)
final_coverage_file = bedgraph(out_sorted_bam)
stats(out_sorted_bam)
ptr(final_coverage_file)
<<<<<<< HEAD:ptr.py
=======
>>>>>>> 82a9fde8f5c26edb4007075eb056584348a71e72:pipeline.py
elif steps_list[0] == "ptr":
final_coverage_file = "%s/%s_coverage.bed" % (args.output_folder, args.analysis_name)
out_sorted_bam = "%s/%s_aln_sort.bam" % (args.output_folder, args.analysis_name)
stats(out_sorted_bam)
ptr(final_coverage_file)
elif steps_list[0] == "stats":
out_sorted_bam = "%s/%s_aln_sort.bam" % (args.output_folder, args.analysis_name)
stats(out_sorted_bam)
else:
keep_logging(
'Seems like the Analysis Steps are not in sequential order. Please recheck the -steps argument and run the pipeline again',
'Seems like the Analysis Steps are not in sequential order. Please recheck the -steps argument and run the pipeline again',
logger, 'exception')
## Check Subroutines
def usage():
print "Usage: python pipeline.py [-h] -PE1 path-to-forward-PE-read -PE2 path-to-reverse-PE-read -o path-to-OUTPUT_FOLDER -analysis ANALYSIS_NAME -index INDEX_NAME_as_per_config_file \n"
# Validate Filenames for any unsupported characters
def Validate_filename( name ):
pattern_strings = ['\.', '\&', '\>', 'aaa', '\*']
pattern_string = '|'.join(pattern_strings)
searchobj = re.search(pattern_string, name, flags=0)
if searchobj:
print "The file " + name + " contains unsupported characters such as quotes, spaces, or &:%?*><\$. \nPlease Provide another file name.\n"
exit()
def file_exists(path1, path2, reference):
if not os.path.isfile(path1):
file_basename = os.path.basename(path1)
keep_logging('The input file {} does not exists. Please provide another file or check the files path.\n'.format(file_basename), 'The input file {} does not exists. Please provide another file or check the files path.\n'.format(file_basename), logger, 'exception')
exit()
if path2 is not None:
if not os.path.isfile(path2):
file_basename = os.path.basename(path2)
keep_logging('The input file {} does not exists. Please provide another file or check the files path.\n'.format(file_basename), 'The input file {} does not exists. Please provide another file or check the files path.\n'.format(file_basename), logger, 'exception')
exit()
if not os.path.isfile(reference):
file_basename = os.path.basename(reference)
keep_logging('The reference fasta file {} does not exists. Please provide another file or check the files path.\n'.format(file_basename), 'The reference fasta file {} does not exists. Please provide another file or check the files path.\n'.format(file_basename), logger, 'exception')
exit()
if ConfigSectionMap("pipeline", Config)['aligner'] == "bwa":
ref_index_suffix1 = reference + ".bwt"
ref_index_suffix2 = reference + ".amb"
ref_index_suffix3 = reference + ".ann"
ref_index_suffix4 = reference + ".sa"
ref_index_suffix5 = reference + ".pac"
elif ConfigSectionMap("pipeline", Config)['aligner'] == "bowtie":
ref_index_suffix1 = reference + ".1.bt2"
ref_index_suffix2 = reference + ".2.bt2"
ref_index_suffix3 = reference + ".3.bt2"
ref_index_suffix4 = reference + ".4.ebwt"
ref_index_suffix5 = reference + ".rev.1.bt2"
ref_index_suffix6 = reference + ".rev.2.bt2"
else:
print "Please change the aligner section in config file."
print "Different Aligner in config file"
if not os.path.isfile(ref_index_suffix1):
# file_basename = os.path.basename(reference)
keep_logging('The reference index files given below does not exists:\n {}\n {}\n {}\n {}\n {}'.format(ref_index_suffix1, ref_index_suffix2, ref_index_suffix3, ref_index_suffix4, ref_index_suffix5), 'The reference index files given below does not exists:\n {}\n {}\n {}\n {}\n {}'.format(ref_index_suffix1, ref_index_suffix2, ref_index_suffix3, ref_index_suffix4, ref_index_suffix5), logger, 'warning')
create_index(reference, ref_index_suffix1, ref_index_suffix2, ref_index_suffix3, ref_index_suffix4, ref_index_suffix5)
else:
keep_logging('Index file already exists.', 'Index file already exists.', logger, 'info')
ref_fai_index = reference + ".fai"
if not os.path.isfile(ref_fai_index):
# file_basename = os.path.basename(reference)
keep_logging('The reference fai index file {} required for samtools does not exists.'.format(ref_fai_index), 'The reference fai index file {} required for samtools does not exists.'.format(ref_fai_index), logger, 'warning')
create_fai_index(reference, ref_fai_index)
else:
keep_logging('Samtools fai Index file already exists.', 'Samtools fai Index file already exists.', logger, 'info')
dict_name = os.path.splitext(os.path.basename(reference))[0] + ".dict"
if not os.path.isfile(ConfigSectionMap(args.index, Config)['ref_path'] + "/" + dict_name):
keep_logging('The reference seq dict file {} required for GATK and PICARD does not exists.'.format(dict_name), 'The reference seq dict file {} required for GATK and PICARD does not exists.'.format(dict_name), logger, 'warning')
picard_seqdict(dict_name, reference)
else:
keep_logging('The reference seq dict file required for GATK and PICARD exists.', 'The reference seq dict file required for GATK and PICARD exists.', logger, 'info')
def java_check():
keep_logging('Checking Java Availability...', 'Checking Java Availability...', logger, 'info')
jd = sp.check_output(["java", "-version"], stderr=sp.STDOUT)
jd_version = jd.split('\n', 1)[0]
if len(jd) < 1:
keep_logging('Unable to find a java runtime environment. The pipeline requires java 6 or later.', 'Unable to find a java runtime environment. The pipeline requires java 6 or later.', logger, 'exception')
else:
keep_logging('Java Availability Check completed ...{}'.format(jd_version), 'Java Availability Check completed ...{}'.format(jd_version), logger, 'info')
def fileformat(file1, file2, final_out):
print "Checking File format....\n"
if not file1.endswith('.fastq.gz'):
base = os.path.basename(file1)
os.path.splitext(base)
file_1 = os.path.splitext(base)[0]
cmdstring = "gzip -d " + file1 + " > " + final_out + file_1
print "Compressing input file " + base
os.system(cmdstring)
if not file2.endswith('.fastq.gz'):
base = os.path.basename(file2)
os.path.splitext(base)
file_2 = os.path.splitext(base)[0]
cmdstring = "gzip -d " + file2 + " > " + final_out + file_2
print "Compressing input file " + base
os.system(cmdstring)
def make_sure_path_exists(out_path):
try:
os.makedirs(out_path)
except OSError as exception:
if exception.errno != errno.EEXIST:
keep_logging('Errors in output folder path! please change the output path or analysis name.', 'Errors in output folder path! please change the output path or analysis name', logger, 'exception')
exit()
def create_index(reference,ref_index_suffix1, ref_index_suffix2, ref_index_suffix3, ref_index_suffix4, ref_index_suffix5):
aligner = ConfigSectionMap("pipeline", Config)['aligner']
keep_logging('Creating Index of reference fasta file for {} aligner.'.format(aligner), 'Creating Index of reference fasta file for {} aligner'.format(aligner), logger, 'info')
if aligner == "bwa":
cmd = "%s %s %s" % (ConfigSectionMap("bwa", Config)['base_cmd'], ConfigSectionMap("bwa", Config)['index'], reference)
keep_logging(cmd, cmd, logger, 'debug')
try:
call(cmd, logger)
except sp.CalledProcessError:
keep_logging('Error in {} Indexer. Exiting.'.format(aligner), 'Error in {} Indexer. Exiting.'.format(aligner), logger, 'exception')
sys.exit(1)
if not os.path.isfile(ref_index_suffix1):
keep_logging('The {} reference index files were not created properly. Please try to create the index files again or manually.'.format(aligner), 'The {} reference index files were not created properly. Please try to create the index files again or manually.'.format(aligner), logger, 'exception')
elif aligner == "bowtie":
cmd = "%s/%s/%s %s %s" % (ConfigSectionMap("bin_path", Config)['binbase'], ConfigSectionMap("bowtie", Config)['bowtie_bin'], ConfigSectionMap("bowtie", Config)['build_cmd'], reference, reference)
keep_logging(cmd, cmd, logger, 'debug')
try:
call(cmd, logger)
except sp.CalledProcessError:
keep_logging('Error in {} Indexer. Exiting.'.format(aligner), 'Error in {} Indexer. Exiting.'.format(aligner), logger, 'exception')
sys.exit(1)
if not os.path.isfile(ref_index_suffix1):
keep_logging('The {} reference index files were not created properly. Please try to create the index files again or manually.'.format(aligner), 'The {} reference index files were not created properly. Please try to create the index files again or manually.'.format(aligner), logger, 'exception')
else:
print "Different Aligner in config file"
def create_fai_index(reference, ref_fai_index):
keep_logging('Creating FAI Index using Samtools.', 'Creating FAI Index using Samtools.', logger, 'info')
cmd = "%s %s %s" % (ConfigSectionMap("samtools", Config)['base_cmd'], ConfigSectionMap("samtools", Config)['faiindex'], reference)
keep_logging(cmd, cmd, logger, 'debug')
try:
call(cmd, logger)
except sp.CalledProcessError:
keep_logging('Error in Samtools FAI Indexing step. Exiting.', 'Error in Samtools FAI Indexing step. Exiting.', logger, 'exception')
sys.exit(1)
if not os.path.isfile(ref_fai_index):
keep_logging('The reference fai index file {} was not created properly.\n Please try to create the samtools fai index files manually. \n'.format(ref_fai_index), 'The reference fai index file {} was not created properly.\n Please try to create the samtools fai index files manually. \n'.format(ref_fai_index), logger, 'exception')
else:
keep_logging('Samtools Fai Index file created.', 'Samtools Fai Index file created.', logger, 'info')
def picard_seqdict(dict_name, reference):
#dict_name = os.path.splitext(os.path.basename(reference_filename))[0] + ".dict"
keep_logging('Creating Sequence Dictionary using Picard.', 'Creating Sequence Dictionary using Picard.', logger, 'info')
cmd = "java -jar %s/%s/%s CreateSequenceDictionary REFERENCE=%s OUTPUT=%s/%s" % (ConfigSectionMap("bin_path", Config)['binbase'], ConfigSectionMap("picard", Config)['picard_bin'], ConfigSectionMap("picard", Config)['base_cmd'], reference, ConfigSectionMap(args.index, Config)['ref_path'], dict_name)
keep_logging(cmd, cmd, logger, 'debug')
try:
call(cmd, logger)
except sp.CalledProcessError:
keep_logging('Error in Picard Sequence Dictionary creation step. Exiting.', 'Error in Picard Sequence Dictionary creation step. Exiting.', logger, 'exception')
sys.exit(1)
""" Start of Main Method/Pipeline """
if __name__ == '__main__':
# Set up logging modules and config file
start_time = datetime.now().strftime('%Y-%m-%d %H:%M:%S')
start_time_2 = datetime.now()
args = parser().parse_args()
global config_file
global log_unique_time
global logger
global files_to_delete
files_to_delete = []
if args.config:
config_file = args.config
else:
config_file = os.path.dirname(os.path.abspath(__file__)) + "/config"
if args.output_folder != '':
args.output_folder += '/'
make_sure_path_exists(args.output_folder)
log_unique_time = datetime.now().strftime('%Y_%m_%d_%H_%M_%S')
logger = generate_logger(args.output_folder, args.analysis_name, log_unique_time)
global Config
Config = ConfigParser.ConfigParser()
Config.read(config_file)
keep_logging('START: Pipeline', 'START: Pipeline', logger, 'info')
pipeline(args, logger)
keep_logging('END: Pipeline', 'End: Pipeline', logger, 'info')
time_taken = datetime.now() - start_time_2
keep_logging('Total Time taken: {}'.format(time_taken), 'Total Time taken: {}'.format(time_taken), logger, 'info')