def _remove_self_tagdust(tagdust_config, input_files):
""" creates a temporary contaminant fasta file for each input file which
does not have its own sequence in it, for filtering with tagdust
"""
short_names = map(_short_name, input_files)
contam = tagdust_config["contaminants"]
def suffix_not_in_name(seq, suffix):
return not suffix in seq.id
# build the predicate to test if the short name of the input file is
# in the fasta sequence
predicates = [partial(suffix_not_in_name, suffix=x) for x in short_names]
# filter the contaminant fasta file, outputting a separate file for
# each input file
filtered_files = [filter_seqio(contam, predicate, suffix)
for suffix, predicate in zip(short_names, predicates)]
return filtered_files
def _remove_self_tagdust(tagdust_config, input_files):
""" creates a temporary contaminant fasta file for each input file which
does not have its own sequence in it, for filtering with tagdust
"""
short_names = map(_short_name, input_files)
contam = tagdust_config["contaminants"]
def suffix_not_in_name(seq, suffix):
return not suffix in seq.id
# build the predicate to test if the short name of the input file is
# in the fasta sequence
predicates = [partial(suffix_not_in_name, suffix=x) for x in short_names]
# filter the contaminant fasta file, outputting a separate file for
# each input file
filtered_files = [
filter_seqio(contam, predicate, suffix)
for suffix, predicate in zip(short_names, predicates)
]
return filtered_files
def main(config_file):
with open(config_file) as in_handle:
config = yaml.load(in_handle)
setup_logging(config)
start_cluster(config)
# after the cluster is up, import the view to i
from bipy.cluster import view
input_files = config["input"]
results_dir = config["dir"]["results"]
# make the needed directories
map(safe_makedir, config["dir"].values())
curr_files = input_files
## qc steps
for stage in config["run"]:
if stage == "fastqc":
# run the basic fastqc
logger.info("Running %s on %s" % (stage, str(curr_files)))
fastqc_config = config["stage"][stage]
fastqc_outputs = view.map(fastqc.run, curr_files,
[fastqc_config] * len(curr_files),
[config] * len(curr_files))
# this does nothing for now, not implemented yet
summary_file = _combine_fastqc(fastqc_outputs)
if stage == "trim":
logger.info("Trimming poor quality ends "
" from %s" % (str(curr_files)))
nlen = len(curr_files)
min_length = str(config["stage"][stage].get("min_length", 20))
# trim low quality ends of reads
# do this dirty for now
out_dir = os.path.join(results_dir, "trimmed")
safe_makedir(out_dir)
out_files = [append_stem(os.path.basename(x), "trim") for
x in curr_files]
out_files = [os.path.join(out_dir, x) for x in out_files]
# XXX remove the magic number of 10 the length of the
# minimum read to keep
out_files = view.map(sickle.run, curr_files,
["se"] * nlen,
["sanger"] * nlen,
[min_length] * nlen,
out_files)
curr_files = out_files
if stage == "tagdust":
input_files = curr_files
# remove tags matching the other miRNA tested
logger.info("Running %s on %s." % (stage, input_files))
tagdust_config = config["stage"][stage]
tagdust_outputs = view.map(tagdust.run, input_files,
[tagdust_config] * len(input_files),
[config] * len(input_files))
curr_files = [x[0] for x in tagdust_outputs]
if stage == "filter_length":
# filter out reads below or above a certain length
filter_config = config["stage"][stage]
min_length = filter_config.get("min_length", 0)
max_length = filter_config.get("max_length", MAX_READ_LENGTH)
# length predicate
def length_filter(x):
return min_length < len(x.seq) < max_length
# filter the input reads based on length
# parallelizing this doesn't seem to work
# ipython can't accept closures as an argument to view.map()
"""
filtered_fastq = view.map(filter_seqio, tagdust_outputs,
[lf] * len(tagdust_outputs),
["filt"] * len(tagdust_outputs),
["fastq"] * len(tagdust_outputs))"""
out_files = [append_stem(os.path.basename(input_file[0]),
"filt") for input_file in tagdust_outputs]
out_dir = os.path.join(config["dir"]["results"],
"length_filtered")
safe_makedir(out_dir)
out_files = [os.path.join(out_dir, x) for x in out_files]
filtered_fastq = [filter_seqio(x[0], length_filter, y, "fastq")
for x, y in zip(tagdust_outputs, out_files)]
curr_files = filtered_fastq
if stage == "count_ends":
logger.info("Compiling nucleotide counts at 3' and 5' ends.")
# count the nucleotide at the end of each read
def count_ends(x, y):
""" keeps a running count of an arbitrary set of keys
during the reduce step """
x[y] = x.get(y, 0) + 1
return x
def get_3prime_end(x):
return str(x.seq[-1])
def get_5prime_end(x):
return str(x.seq[0])
def output_counts(end_function, count_file):
# if the count_file already exists, skip
outdir = os.path.join(config["dir"]["results"], stage)
safe_makedir(outdir)
count_file = os.path.join(outdir, count_file)
if os.path.exists(count_file):
return count_file
# outputs a tab file of the counts at the end
# of the fastq files kj
counts = [reduce(count_ends,
apply_seqio(x, end_function, kind="fastq"),
{}) for x in curr_files]
df = pd.DataFrame(counts,
index=map(_short_name, curr_files))
df = df.astype(float)
total = df.sum(axis=1)
df = df.div(total, axis=0)
df["total"] = total
df.to_csv(count_file, sep="\t")
output_counts(get_3prime_end, "3prime_counts.tsv")
output_counts(get_5prime_end, "5prime_counts.tsv")
if stage == "tophat":
tophat_config = config["stage"][stage]
logger.info("Running tophat on %s" % (str(curr_files)))
nlen = len(curr_files)
pair_file = None
ref_file = tophat_config["annotation"]
out_base = os.path.join(results_dir, "mirna")
align_dir = os.path.join(results_dir, "tophat")
config = config
tophat_files = view.map(tophat.align,
curr_files,
[pair_file] * nlen,
[ref_file] * nlen,
[out_base] * nlen,
[align_dir] * nlen,
[config] * nlen)
curr_files = tophat_files
if stage == "novoalign":
logger.info("Running novoalign on %s" % (str(curr_files)))
# align
ref = config["genome"]["file"]
novoalign_config = config["stage"][stage]
aligned_outputs = view.map(novoalign.run, curr_files,
[ref] * len(curr_files),
[novoalign_config] * len(curr_files),
[config] * len(curr_files))
# convert sam to bam, sort and index
picard = BroadRunner(config["program"]["picard"], None, {})
bamfiles = view.map(picardrun.picard_formatconverter,
[picard] * len(aligned_outputs),
aligned_outputs)
sorted_bf = view.map(picardrun.picard_sort,
[picard] * len(bamfiles),
bamfiles)
view.map(picardrun.picard_index, [picard] * len(sorted_bf),
sorted_bf)
# these files are the new starting point for the downstream
# analyses, so copy them over into the data dir and setting
# them to read only
#data_dir = os.path.join(config["dir"]["data"], stage)
#safe_makedir(data_dir)
#view.map(shutil.copy, sorted_bf, [data_dir] * len(sorted_bf))
#new_files = [os.path.join(data_dir, x) for x in
# map(os.path.basename, sorted_bf)]
#[os.chmod(x, stat.S_IREAD | stat.S_IRGRP) for x in new_files]
# index the bam files for later use
#view.map(picardrun.picard_index, [picard] * len(new_files),
# new_files)
curr_files = sorted_bf
if stage == "new_coverage":
logger.info("Calculating RNASeq metrics on %s." % (curr_files))
nrun = len(curr_files)
ref = blastn.prepare_ref_file(config["stage"][stage]["ref"],
config)
ribo = config["stage"][stage]["ribo"]
picard = BroadRunner(config["program"]["picard"], None, {})
out_dir = os.path.join(results_dir, "new_coverage")
safe_makedir(out_dir)
out_files = [replace_suffix(os.path.basename(x),
"metrics") for x in curr_files]
out_files = [os.path.join(out_dir, x) for x in out_files]
out_files = view.map(picardrun.picard_rnaseq_metrics,
[picard] * nrun,
curr_files,
[ref] * nrun,
[ribo] * nrun,
out_files)
curr_files = out_files
if stage == "coverage":
gtf = blastn.prepare_ref_file(config["annotation"], config)
logger.info("Calculating coverage of features in %s for %s"
% (gtf, str(sorted_bf)))
out_files = [replace_suffix(x, "counts.bed") for
x in sorted_bf]
out_dir = os.path.join(results_dir, stage)
safe_makedir(out_dir)
logger.info(out_files)
out_files = [os.path.join(out_dir,
os.path.basename(x)) for x in out_files]
logger.info(out_files)
view.map(bedtools.count_overlaps, sorted_bf,
[gtf] * len(sorted_bf),
out_files)
if stage == "htseq-count":
nfiles = len(curr_files)
htseq_config = _get_stage_config(config, stage)
htseq_outputs = view.map(htseq_count.run_with_config,
aligned_outputs,
[config] * nfiles,
[stage] * nfiles)
column_names = _get_short_names(input_files)
logger.info("Column names: %s" % (column_names))
out_file = os.path.join(config["dir"]["results"], stage,
"combined.counts")
combined_out = htseq_count.combine_counts(htseq_outputs,
column_names,
out_file)
if stage == "bedtools_intersect":
bedfiles = config["stage"]["bedtools_intersect"].get("bed", None)
out_dir = os.path.join(results_dir, stage)
safe_makedir(out_dir)
for bedfile in bedfiles:
bedbase, bedext = os.path.splitext(bedfile)
out_files = [remove_suffix(x) for x in sorted_bf]
out_files = [os.path.join(out_dir, os.path.basename(x)) for x in
out_files]
out_files = ["_vs_".join([x, os.path.basename(bedbase)])
for x in out_files]
out_files = [".".join([x, "bam"]) for x in out_files]
test_out = map(bedtools.intersectbam2bed, sorted_bf,
[bedfile] * len(sorted_bf),
[False] * len(sorted_bf),
out_files)
count_files = [replace_suffix(x, "stats") for x in
out_files]
map(write_ratios, sorted_bf, out_files, count_files)
if stage == "piranha":
piranha_runner = piranha.PiranhaStage(config)
out_files = view.map(piranha_runner, curr_files)
stop_cluster()
def main(config_file):
with open(config_file) as in_handle:
config = yaml.load(in_handle)
setup_logging(config)
start_cluster(config)
# after the cluster is up, import the view to i
from bipy.cluster import view
input_files = config["input"]
results_dir = config["dir"]["results"]
# make the needed directories
map(safe_makedir, config["dir"].values())
curr_files = input_files
## qc steps
for stage in config["run"]:
if stage == "fastqc":
# run the basic fastqc
logger.info("Running %s on %s" % (stage, str(curr_files)))
fastqc_config = config["stage"][stage]
fastqc_outputs = view.map(fastqc.run, curr_files,
[fastqc_config] * len(curr_files),
[config] * len(curr_files))
# this does nothing for now, not implemented yet
summary_file = _combine_fastqc(fastqc_outputs)
if stage == "trim":
logger.info("Trimming poor quality ends "
" from %s" % (str(curr_files)))
nlen = len(curr_files)
min_length = str(config["stage"][stage].get("min_length", 20))
# trim low quality ends of reads
# do this dirty for now
out_dir = os.path.join(results_dir, "trimmed")
safe_makedir(out_dir)
out_files = [
append_stem(os.path.basename(x), "trim") for x in curr_files
]
out_files = [os.path.join(out_dir, x) for x in out_files]
# XXX remove the magic number of 10 the length of the
# minimum read to keep
out_files = view.map(sickle.run, curr_files, ["se"] * nlen,
["sanger"] * nlen, [min_length] * nlen,
out_files)
curr_files = out_files
if stage == "tagdust":
input_files = curr_files
# remove tags matching the other miRNA tested
logger.info("Running %s on %s." % (stage, input_files))
tagdust_config = config["stage"][stage]
tagdust_outputs = view.map(tagdust.run, input_files,
[tagdust_config] * len(input_files),
[config] * len(input_files))
curr_files = [x[0] for x in tagdust_outputs]
if stage == "filter_length":
# filter out reads below or above a certain length
filter_config = config["stage"][stage]
min_length = filter_config.get("min_length", 0)
max_length = filter_config.get("max_length", MAX_READ_LENGTH)
# length predicate
def length_filter(x):
return min_length < len(x.seq) < max_length
# filter the input reads based on length
# parallelizing this doesn't seem to work
# ipython can't accept closures as an argument to view.map()
"""
filtered_fastq = view.map(filter_seqio, tagdust_outputs,
[lf] * len(tagdust_outputs),
["filt"] * len(tagdust_outputs),
["fastq"] * len(tagdust_outputs))"""
out_files = [
append_stem(os.path.basename(input_file[0]), "filt")
for input_file in tagdust_outputs
]
out_dir = os.path.join(config["dir"]["results"], "length_filtered")
safe_makedir(out_dir)
out_files = [os.path.join(out_dir, x) for x in out_files]
filtered_fastq = [
filter_seqio(x[0], length_filter, y, "fastq")
for x, y in zip(tagdust_outputs, out_files)
]
curr_files = filtered_fastq
if stage == "count_ends":
logger.info("Compiling nucleotide counts at 3' and 5' ends.")
# count the nucleotide at the end of each read
def count_ends(x, y):
""" keeps a running count of an arbitrary set of keys
during the reduce step """
x[y] = x.get(y, 0) + 1
return x
def get_3prime_end(x):
return str(x.seq[-1])
def get_5prime_end(x):
return str(x.seq[0])
def output_counts(end_function, count_file):
# if the count_file already exists, skip
outdir = os.path.join(config["dir"]["results"], stage)
safe_makedir(outdir)
count_file = os.path.join(outdir, count_file)
if os.path.exists(count_file):
return count_file
# outputs a tab file of the counts at the end
# of the fastq files kj
counts = [
reduce(count_ends,
apply_seqio(x, end_function, kind="fastq"), {})
for x in curr_files
]
df = pd.DataFrame(counts, index=map(_short_name, curr_files))
df = df.astype(float)
total = df.sum(axis=1)
df = df.div(total, axis=0)
df["total"] = total
df.to_csv(count_file, sep="\t")
output_counts(get_3prime_end, "3prime_counts.tsv")
output_counts(get_5prime_end, "5prime_counts.tsv")
if stage == "tophat":
tophat_config = config["stage"][stage]
logger.info("Running tophat on %s" % (str(curr_files)))
nlen = len(curr_files)
pair_file = None
ref_file = tophat_config["annotation"]
out_base = os.path.join(results_dir, "mirna")
align_dir = os.path.join(results_dir, "tophat")
config = config
tophat_files = view.map(tophat.align, curr_files,
[pair_file] * nlen, [ref_file] * nlen,
[out_base] * nlen, [align_dir] * nlen,
[config] * nlen)
curr_files = tophat_files
if stage == "novoalign":
logger.info("Running novoalign on %s" % (str(curr_files)))
# align
ref = config["genome"]["file"]
novoalign_config = config["stage"][stage]
aligned_outputs = view.map(novoalign.run, curr_files,
[ref] * len(curr_files),
[novoalign_config] * len(curr_files),
[config] * len(curr_files))
# convert sam to bam, sort and index
picard = BroadRunner(config["program"]["picard"], None, {})
bamfiles = view.map(picardrun.picard_formatconverter,
[picard] * len(aligned_outputs),
aligned_outputs)
sorted_bf = view.map(picardrun.picard_sort,
[picard] * len(bamfiles), bamfiles)
view.map(picardrun.picard_index, [picard] * len(sorted_bf),
sorted_bf)
# these files are the new starting point for the downstream
# analyses, so copy them over into the data dir and setting
# them to read only
#data_dir = os.path.join(config["dir"]["data"], stage)
#safe_makedir(data_dir)
#view.map(shutil.copy, sorted_bf, [data_dir] * len(sorted_bf))
#new_files = [os.path.join(data_dir, x) for x in
# map(os.path.basename, sorted_bf)]
#[os.chmod(x, stat.S_IREAD | stat.S_IRGRP) for x in new_files]
# index the bam files for later use
#view.map(picardrun.picard_index, [picard] * len(new_files),
# new_files)
curr_files = sorted_bf
if stage == "new_coverage":
logger.info("Calculating RNASeq metrics on %s." % (curr_files))
nrun = len(curr_files)
ref = blastn.prepare_ref_file(config["stage"][stage]["ref"],
config)
ribo = config["stage"][stage]["ribo"]
picard = BroadRunner(config["program"]["picard"], None, {})
out_dir = os.path.join(results_dir, "new_coverage")
safe_makedir(out_dir)
out_files = [
replace_suffix(os.path.basename(x), "metrics")
for x in curr_files
]
out_files = [os.path.join(out_dir, x) for x in out_files]
out_files = view.map(picardrun.picard_rnaseq_metrics,
[picard] * nrun, curr_files, [ref] * nrun,
[ribo] * nrun, out_files)
curr_files = out_files
if stage == "coverage":
gtf = blastn.prepare_ref_file(config["annotation"], config)
logger.info("Calculating coverage of features in %s for %s" %
(gtf, str(sorted_bf)))
out_files = [replace_suffix(x, "counts.bed") for x in sorted_bf]
out_dir = os.path.join(results_dir, stage)
safe_makedir(out_dir)
logger.info(out_files)
out_files = [
os.path.join(out_dir, os.path.basename(x)) for x in out_files
]
logger.info(out_files)
view.map(bedtools.count_overlaps, sorted_bf,
[gtf] * len(sorted_bf), out_files)
if stage == "htseq-count":
nfiles = len(curr_files)
htseq_config = _get_stage_config(config, stage)
htseq_outputs = view.map(htseq_count.run_with_config,
aligned_outputs, [config] * nfiles,
[stage] * nfiles)
column_names = _get_short_names(input_files)
logger.info("Column names: %s" % (column_names))
out_file = os.path.join(config["dir"]["results"], stage,
"combined.counts")
combined_out = htseq_count.combine_counts(htseq_outputs,
column_names, out_file)
if stage == "bedtools_intersect":
bedfiles = config["stage"]["bedtools_intersect"].get("bed", None)
out_dir = os.path.join(results_dir, stage)
safe_makedir(out_dir)
for bedfile in bedfiles:
bedbase, bedext = os.path.splitext(bedfile)
out_files = [remove_suffix(x) for x in sorted_bf]
out_files = [
os.path.join(out_dir, os.path.basename(x))
for x in out_files
]
out_files = [
"_vs_".join([x, os.path.basename(bedbase)])
for x in out_files
]
out_files = [".".join([x, "bam"]) for x in out_files]
test_out = map(bedtools.intersectbam2bed, sorted_bf,
[bedfile] * len(sorted_bf),
[False] * len(sorted_bf), out_files)
count_files = [replace_suffix(x, "stats") for x in out_files]
map(write_ratios, sorted_bf, out_files, count_files)
if stage == "piranha":
piranha_runner = piranha.PiranhaStage(config)
out_files = view.map(piranha_runner, curr_files)
stop_cluster()