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/
three_p_experiment.py
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three_p_experiment.py
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import matplotlib
matplotlib.use('Agg', warn=False)
import os
import positions
import visualize
import trim
import pysam
from Sequencing import fastq, sam, mapping_tools, genomes, utilities
from Sequencing.Parallel import map_reduce
import Serialize.read_positions
from Sequencing.Serialize import array_1d
import rna_experiment
from Sequencing.annotation import Annotation_factory
from collections import Counter
class ThreePExperiment(rna_experiment.RNAExperiment):
num_stages = 2
specific_results_files = [
('trimmed_reads', 'fastq', '{name}_trimmed.fastq'),
('too_short_lengths', array_1d, '{name}_too_short_lengths.txt'),
('trimmed_lengths', array_1d, '{name}_trimmed_lengths.txt'),
('nongenomic_lengths', array_1d, '{name}_nongenomic_lengths.txt'),
('extended', 'bam', '{name}_extended.bam'),
('extended_filtered', 'bam', '{name}_extended_filtered.bam'),
]
specific_figure_files = []
specific_outputs = [
['trimmed_lengths',
'too_short_lengths',
'extended',
'extended_filtered',
'nongenomic_lengths',
],
['metagene_positions',
'read_positions',
],
]
specific_work = [
['preprocess',
'map_tophat',
'extend_mappings',
'filter_mappings',
],
['get_polyA_positions',
'get_metagene_positions',
],
]
specific_cleanup = [
[],
['plot_starts_and_ends',
],
]
def __init__(self, **kwargs):
super(ThreePExperiment, self).__init__(**kwargs)
self.max_read_length = self.get_max_read_length()
self.min_length = 12
self.trim_function = trim.bound_trim['polyA']
def preprocess(self):
reads = self.get_reads()
trimmed_reads = self.trim_reads(reads)
with open(self.file_names['trimmed_reads'], 'w') as trimmed_fh:
for read in trimmed_reads:
trimmed_fh.write(str(read))
def map_tophat(self):
mapping_tools.map_tophat([self.file_names['trimmed_reads']],
self.file_names['bowtie2_index_prefix'],
self.file_names['genes'],
self.file_names['transcriptome_index'],
self.file_names['tophat_dir'],
no_sort=True,
)
def extend_mappings(self):
trim.extend_polyA_ends(self.file_names['accepted_hits'],
self.file_names['extended'],
self.file_names['genome'],
)
def filter_mappings(self):
num_unmapped = 0
num_entirely_genomic = 0
num_nonunique = 0
num_unique = 0
nongenomic_lengths = Counter()
sam_file = pysam.Samfile(self.file_names['accepted_hits'])
region_fetcher = genomes.build_region_fetcher(self.file_names['genome'],
load_references=True,
sam_file=sam_file,
)
extended_sorter = sam.AlignmentSorter(sam_file.references,
sam_file.lengths,
self.file_names['extended'],
)
filtered_sorter = sam.AlignmentSorter(sam_file.references,
sam_file.lengths,
self.file_names['extended_filtered'],
)
extended_mappings = (trim.extend_polyA_end(mapping, region_fetcher) for mapping in sam_file)
mapping_groups = utilities.group_by(extended_mappings, lambda m: m.qname)
with extended_sorter, filtered_sorter:
for qname, group in mapping_groups:
for m in group:
extended_sorter.write(m)
min_nongenomic_length = min(trim.get_nongenomic_length(m) for m in group)
nongenomic_lengths[min_nongenomic_length] += 1
if min_nongenomic_length == 0:
num_entirely_genomic += 1
continue
nonunique = len(group) > 1 or any(m.mapq < 40 for m in group)
if nonunique:
num_nonunique += 1
continue
num_unique += 1
for m in group:
filtered_sorter.write(m)
self.summary.extend(
[('Mapped with no non-genomic A\'s', num_entirely_genomic),
('Nonunique', num_nonunique),
('Unique', num_unique),
],
)
nongenomic_lengths = utilities.counts_to_array(nongenomic_lengths)
self.write_file('nongenomic_lengths', nongenomic_lengths)
def get_polyA_positions(self):
piece_CDSs, max_gene_length = self.get_CDSs()
gene_infos = positions.get_Transcript_position_counts(self.merged_file_names['extended'],
piece_CDSs,
[],
left_buffer=500,
right_buffer=500,
)
self.read_positions = {name: info['three_prime_positions']
for name, info in gene_infos.iteritems()}
self.write_file('read_positions', self.read_positions)
def get_metagene_positions(self):
piece_CDSs, max_gene_length = self.get_CDSs()
read_positions = self.load_read_positions()
processed_read_positions = {}
for name in read_positions:
gene = {'three_prime_genomic': read_positions[name][0],
'three_prime_nongenomic': read_positions[name]['all'] - read_positions[name][0],
'three_prime_nonunique': three_prime_counts['all_nonunique'],
'sequence': read_positions[name]['sequence'],
}
processed_read_positions[name] = gene
metagene_positions = positions.compute_metagene_positions(piece_CDSs,
processed_read_positions,
max_gene_length,
)
self.write_file('metagene_positions', metagene_positions)
def plot_starts_and_ends(self):
metagene_positions = self.read_file('metagene_positions')
visualize.plot_metagene_positions(metagene_positions,
self.figure_file_names['starts_and_ends'],
['three_prime_genomic', 'three_prime_nongenomic'],
)
def get_total_eligible_reads(self):
summary_pairs = self.read_file('summary')
summary_dict = {name: values[0] for name, values in summary_pairs}
total_mapped_reads = summary_dict['Nonunique'] + summary_dict['Unique']
return total_mapped_reads
if __name__ == '__main__':
script_path = os.path.realpath(__file__)
map_reduce.controller(ThreePExperiment, script_path)