def _goleft_indexcov(bam_file, data, out_dir):
"""Use goleft indexcov to estimate coverage distributions using BAM index.
Only used for whole genome runs as captures typically don't have enough data
to be useful for index-only summaries.
"""
if not dd.get_coverage_interval(data) == "genome":
return []
out_dir = utils.safe_makedir(os.path.join(out_dir, "indexcov"))
out_files = [os.path.join(out_dir, "%s-indexcov.%s" % (dd.get_sample_name(data), ext))
for ext in ["roc", "ped", "bed.gz"]]
if not utils.file_uptodate(out_files[-1], bam_file):
with transaction.tx_tmpdir(data) as tmp_dir:
tmp_dir = utils.safe_makedir(os.path.join(tmp_dir, dd.get_sample_name(data)))
gender_chroms = [x.name for x in ref.file_contigs(dd.get_ref_file(data)) if chromhacks.is_sex(x.name)]
gender_args = "--sex %s" % (",".join(gender_chroms)) if gender_chroms else ""
cmd = "goleft indexcov --directory {tmp_dir} {gender_args} -- {bam_file}"
try:
do.run(cmd.format(**locals()), "QC: goleft indexcov")
except subprocess.CalledProcessError as msg:
if not ("indexcov: no usable" in str(msg) or
("indexcov: expected" in str(msg) and "sex chromosomes, found:" in str(msg))):
raise
for out_file in out_files:
orig_file = os.path.join(tmp_dir, os.path.basename(out_file))
if utils.file_exists(orig_file):
utils.copy_plus(orig_file, out_file)
# MultiQC needs non-gzipped/BED inputs so unpack the file
out_bed = out_files[-1].replace(".bed.gz", ".tsv")
if utils.file_exists(out_files[-1]) and not utils.file_exists(out_bed):
with transaction.file_transaction(data, out_bed) as tx_out_bed:
cmd = "gunzip -c %s > %s" % (out_files[-1], tx_out_bed)
do.run(cmd, "Unpack indexcov BED file")
out_files[-1] = out_bed
return [x for x in out_files if utils.file_exists(x)]
def _goleft_indexcov(bam_file, data, out_dir):
"""Use goleft indexcov to estimate coverage distributions using BAM index.
Only used for whole genome runs as captures typically don't have enough data
to be useful for index-only summaries.
"""
if not dd.get_coverage_interval(data) == "genome":
return []
out_dir = utils.safe_makedir(os.path.join(out_dir, "indexcov"))
out_files = [os.path.join(out_dir, "%s-indexcov.%s" % (dd.get_sample_name(data), ext))
for ext in ["roc", "ped", "bed.gz"]]
if not utils.file_uptodate(out_files[-1], bam_file):
with transaction.tx_tmpdir(data) as tmp_dir:
tmp_dir = utils.safe_makedir(os.path.join(tmp_dir, dd.get_sample_name(data)))
gender_chroms = [x.name for x in ref.file_contigs(dd.get_ref_file(data)) if chromhacks.is_sex(x.name)]
gender_args = "--sex %s" % (",".join(gender_chroms)) if gender_chroms else ""
cmd = "goleft indexcov --directory {tmp_dir} {gender_args} -- {bam_file}"
try:
do.run(cmd.format(**locals()), "QC: goleft indexcov")
except subprocess.CalledProcessError as msg:
if not ("indexcov: no usable" in str(msg) or
("indexcov: expected" in str(msg) and "sex chromosomes, found:" in str(msg))):
raise
for out_file in out_files:
orig_file = os.path.join(tmp_dir, os.path.basename(out_file))
if utils.file_exists(orig_file):
utils.copy_plus(orig_file, out_file)
# MultiQC needs non-gzipped/BED inputs so unpack the file
out_bed = out_files[-1].replace(".bed.gz", ".tsv")
if utils.file_exists(out_files[-1]) and not utils.file_exists(out_bed):
with transaction.file_transaction(data, out_bed) as tx_out_bed:
cmd = "gunzip -c %s > %s" % (out_files[-1], tx_out_bed)
do.run(cmd, "Unpack indexcov BED file")
out_files[-1] = out_bed
return [x for x in out_files if utils.file_exists(x)]
def run(bam_file, data, out_dir):
"""Run coverage QC analysis
"""
out = dict()
out_dir = utils.safe_makedir(out_dir)
if dd.get_coverage(data) and dd.get_coverage(data) not in ["None"]:
merged_bed_file = bedutils.clean_file(dd.get_coverage_merged(data),
data,
prefix="cov-",
simple=True)
target_name = "coverage"
elif dd.get_coverage_interval(data) != "genome":
merged_bed_file = dd.get_variant_regions_merged(data)
target_name = "variant_regions"
else:
merged_bed_file = None
target_name = "genome"
avg_depth = cov.get_average_coverage(target_name, merged_bed_file, data)
if target_name == "coverage":
out_files = cov.coverage_region_detailed_stats(target_name,
merged_bed_file, data,
out_dir)
else:
out_files = []
out['Avg_coverage'] = avg_depth
samtools_stats_dir = os.path.join(out_dir, os.path.pardir, 'samtools')
from bcbio.qc import samtools
samtools_stats = samtools.run(bam_file, data,
samtools_stats_dir)["metrics"]
out["Total_reads"] = total_reads = int(samtools_stats["Total_reads"])
out["Mapped_reads"] = mapped = int(samtools_stats["Mapped_reads"])
out["Mapped_paired_reads"] = int(samtools_stats["Mapped_paired_reads"])
out['Duplicates'] = dups = int(samtools_stats["Duplicates"])
if total_reads:
out["Mapped_reads_pct"] = 100.0 * mapped / total_reads
if mapped:
out['Duplicates_pct'] = 100.0 * dups / mapped
if dd.get_coverage_interval(data) == "genome":
mapped_unique = mapped - dups
else:
mapped_unique = readstats.number_of_mapped_reads(data,
bam_file,
keep_dups=False)
out['Mapped_unique_reads'] = mapped_unique
if merged_bed_file:
ontarget = readstats.number_of_mapped_reads(data,
bam_file,
keep_dups=False,
bed_file=merged_bed_file,
target_name=target_name)
out["Ontarget_unique_reads"] = ontarget
if mapped_unique:
out["Ontarget_pct"] = 100.0 * ontarget / mapped_unique
out['Offtarget_pct'] = 100.0 * (mapped_unique -
ontarget) / mapped_unique
if dd.get_coverage_interval(data) != "genome":
# Skip padded calculation for WGS even if the "coverage" file is specified
# the padded statistic makes only sense for exomes and panels
padded_bed_file = bedutils.get_padded_bed_file(
out_dir, merged_bed_file, 200, data)
ontarget_padded = readstats.number_of_mapped_reads(
data,
bam_file,
keep_dups=False,
bed_file=padded_bed_file,
target_name=target_name + "_padded")
out["Ontarget_padded_pct"] = 100.0 * ontarget_padded / mapped_unique
if total_reads:
out['Usable_pct'] = 100.0 * ontarget / total_reads
indexcov_files = _goleft_indexcov(bam_file, data, out_dir)
out_files += [x for x in indexcov_files if x and utils.file_exists(x)]
out = {"metrics": out}
if len(out_files) > 0:
out["base"] = out_files[0]
out["secondary"] = out_files[1:]
return out
def run(bam_file, data, out_dir):
"""Run coverage QC analysis
"""
out = dict()
if dd.get_coverage(data) and dd.get_coverage(data) not in ["None"]:
merged_bed_file = dd.get_coverage_merged(data)
target_name = "coverage"
elif dd.get_coverage_interval(data) != "genome":
merged_bed_file = dd.get_variant_regions_merged(data)
target_name = "variant_regions"
else:
merged_bed_file = None
target_name = "genome"
avg_depth = cov.get_average_coverage(data, bam_file, merged_bed_file, target_name)
out['Avg_coverage'] = avg_depth
samtools_stats_dir = os.path.join(out_dir, os.path.pardir, 'samtools')
from bcbio.qc import samtools
samtools_stats = samtools.run(bam_file, data, samtools_stats_dir)
out["Total_reads"] = total_reads = int(samtools_stats["Total_reads"])
out["Mapped_reads"] = mapped = int(samtools_stats["Mapped_reads"])
out["Mapped_paired_reads"] = int(samtools_stats["Mapped_paired_reads"])
out['Duplicates'] = dups = int(samtools_stats["Duplicates"])
if total_reads:
out["Mapped_reads_pct"] = 100.0 * mapped / total_reads
if mapped:
out['Duplicates_pct'] = 100.0 * dups / mapped
if dd.get_coverage_interval(data) == "genome":
mapped_unique = mapped - dups
else:
mapped_unique = sambamba.number_of_mapped_reads(data, bam_file, keep_dups=False)
out['Mapped_unique_reads'] = mapped_unique
if merged_bed_file:
ontarget = sambamba.number_of_mapped_reads(
data, bam_file, keep_dups=False, bed_file=merged_bed_file, target_name=target_name)
out["Ontarget_unique_reads"] = ontarget
if mapped_unique:
out["Ontarget_pct"] = 100.0 * ontarget / mapped_unique
out['Offtarget_pct'] = 100.0 * (mapped_unique - ontarget) / mapped_unique
if dd.get_coverage_interval(data) != "genome":
# Skip padded calculation for WGS even if the "coverage" file is specified
# the padded statistic makes only sense for exomes and panels
padded_bed_file = bedutils.get_padded_bed_file(out_dir, merged_bed_file, 200, data)
ontarget_padded = sambamba.number_of_mapped_reads(
data, bam_file, keep_dups=False, bed_file=padded_bed_file, target_name=target_name + "_padded")
out["Ontarget_padded_pct"] = 100.0 * ontarget_padded / mapped_unique
if total_reads:
out['Usable_pct'] = 100.0 * ontarget / total_reads
out_files = cov.coverage_region_detailed_stats(data, out_dir,
extra_cutoffs=set([max(1, int(avg_depth * 0.8))]))
for ext in ["coverage.bed", "summary.bed"]:
out_files += [x for x in glob.glob(os.path.join(out_dir, "*%s" % ext)) if os.path.isfile(x)]
indexcov_files = _goleft_indexcov(bam_file, data, out_dir)
out_files += [x for x in indexcov_files if x and utils.file_exists(x)]
out = {"metrics": out}
if len(out_files) > 0:
out["base"] = out_files[0]
out["secondary"] = out_files[1:]
return out
