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#!/usr/bin/python
import sys, argparse, re
from collections import defaultdict, namedtuple, Counter
import pysam
import pbsuite.honey.TGraf as tails
import pbsuite.honey.HSpots as spots
from pbsuite.utils.setupLogging import *
vtypes = ['INS', 'DEL', 'MIS', 'UNK', 'CTX', 'ITX']
USAGE = """
Checks if there are any reads in the given .bam that support predicted SVs
Takes a .bed with the first 6 columns being:
chrom start end name svtype size
- svtype must be one of DEL, INS, MIS
- size is what the SV's size is estimated to be.
If you have single breakpoint events (such as translocations) specify --bedPE
Your input.bed's 9 columns become:
chrom1 start1 end1 orient1 chrom2 start2 end2 orient2 name svtype size
- orient is the directionality of the sequence leading upto the breakpoint (+/-)
RegionBuffer is the +- space in which you consider reads for support
around predicted sv
SizeBuffer is the +- percent of predicted size the read needs to support the SV
Results are an extra column appended to the end of in the format REF[TAILS|SPOTS]
REF:
True if we found evidence of the reference over the region
False if we had the opportunity to support the reference, but didn't.
? if we didn't have the opportunity to support the reference
TAILS/SPOTS:
A comma-separated list of chrBPSchr:start-end(svtype)size*cnt coordinates for
reads that have interrupted or discordant mapping support of the SV.
TAILS/SPOTS:
chr The chromosome
BPS Breakpoint string showing orientations
start/end The breakpoints start and end coordinates
svtype One of %s
cnt Number of reads that support this
""" % ("/".join(vtypes))
class Variant():
def __init__(self, chrom, start, end, svtype, size, read=None):
self.chrom = chrom
self.start = start
self.end = end
self.svtype = svtype
self.size = size
self.read = read
def __str__(self):
return "%s:%d-%d(%s)%d" % (self.chrom, self.start, self.end, self.svtype, self.size)
class BedEntry():
def __init__(self, chrom, start, end, name, svtype, size, *args):
self.chrom = chrom
self.start = int(start)
self.end = int(end)
self.name = name
self.svtype = svtype
self.size = int(size)
self.rest = args
if self.start > self.end:
self.start, self.end = self.end, self.start
def __str__(self):
return "\t".join([str(x) for x in [self.chrom, self.start, \
self.end, self.name, \
self.svtype, self.size, \
"\t".join(self.rest)]])
def __repr__(self):
return "<BedEntry '%s'>" % (str(self).replace('\t',' '))
class BedPEEntry():
def __init__(self, chrom1, start1, end1, orient1, chrom2, start2, end2, orient2, name, svtype, size, *args):
self.chrom1 = chrom1
self.start1 = int(start1)
self.end1 = int(end1)
self.orient1 = orient1
self.chrom2 = chrom2
self.start2 = int(start2)
self.end2 = int(end2)
self.orient2 = orient2
self.name = name
self.svtype = svtype
self.size = int(size)
self.rest = args
def __str__(self):
return "\t".join([str(x) for x in [self.chrom1, self.start1, \
self.end1, self.orient1, \
self.chrom2, self.start2, \
self.end2, self.orient2, \
self.name, self.svtype, \
self.size, "\t".join(self.rest)]])
def __repr__(self):
return "<BedPEEntry '%s'>" % (str(self).replace('\t',' '))
def parseArgs(args):
parser = argparse.ArgumentParser(prog="Honey.py force", description=USAGE, \
formatter_class=argparse.RawDescriptionHelpFormatter)
parser.add_argument("bam", metavar="BAM", type=str, \
help="Assembled Contigs Bam")
parser.add_argument("bed", metavar="BED", type=str, \
help="Bed of locations to force SV Calls")
parser.add_argument("-s", "--sizebuffer", type=float, default=0.35, \
help=("Buffer of estimated sv size to "
"create match (%(default)s)"))
parser.add_argument("-d", "--maxDelta", type=int, default=500, \
help="Max distance between predicted and discovered variant (%(default)s)")
parser.add_argument("-f", "--fetchbuffer", type=int, default=1000, \
help="Buffer for fetching reads from .bam (%(default)s)")
#parser.add_argument("-o", "--overlapbuffer", type=float, default=0.50, \
#help="Percent overlap required from calls to tails (%(default)s)")
parser.add_argument("-q", "--minMapq", type=int, default=100, \
help="Minimum mapping quality of a read and it's tail to consider (%(default)s)")
parser.add_argument("-m", "--minErr", type=int, default=5, \
help="Minimum ins/del error size to consider (%(default)s)")
#parser.add_argument("-a", "--asm", action="store_true", \
#help="Input reads are high-quality contigs")
parser.add_argument("-p", "--bedPE", action="store_true", \
help="Input bed file is bedPE - only tails searching will be performed")
parser.add_argument("--debug", action="store_true", \
help="Verbose logging")
args = parser.parse_args(args)
setupLogging(args.debug)
return args
#MonkeyWrenching of sorts
class FakeBread(tails.Bread):
def __init__(self, svtype, refKey, uRef, dRef, uBreak, dBreak, uDir, \
dDir, is_reverse, uTail, dTail, read):
self.svtype = svtype
self.refKey = refKey
self.uRef = uRef
self.dRef = dRef
self.uBreak = uBreak
self.dBreak = dBreak
self.uDir = uDir
self.dDir = dDir
self.is_reverse = is_reverse
self.uTail = uTail
self.dTail = dTail
self.read = read
self.isInverted = False # This is a problem
self.remainSeq = 1000
def annotate(self):
return tails.Bread.annotate(self)
Fr = namedtuple("fakeread", "is_reverse")
def bedpeToFakey(bed):
"""
Make a fakeBread for PE bed entry
if bed.svtype is unk, we have to make all orientations
"""
# These are all the element I'll be expected to make
# refKey, uBreak, dBreak, uDir, dDir, is_reverse, uTail, dTail
uRef = bed.chrom1; dRef = bed.chrom2
j = [uRef, dRef]; j.sort(); refKey = "_".join(j)
ret = []
if bed.svtype == 'UNK':
#Need to make all orientations
#3i 3e +/-
#+/- to pick the breakpoint locations
bp1 = bed.start1 if True else bed.end1
bp2 = bed.start2 if True else bed.end2
#choose orientation of the first read and automatically flip the second
orient1 = '5' if bed.orient1 == '-' else '3'
orient2 = '5' if bed.orient2 == '+' else '3'
j = [(bp1, orient1, bed.chrom1), (bp2, orient2, bed.chrom2)]; j.sort(); uBP1, dBP2 = j
uBreak, uDir, uRef = uBP1
dBreak, dDir, dRef = dBP2
j = [uRef, dRef]; j.sort(); refKey = "_".join(j)
is_reverse = bed.orient1 == bed.orient2
fr = Fr(False)
ret.append(FakeBread(bed.svtype, refKey, uRef, dRef, uBreak, dBreak, uDir, dDir, is_reverse, 'i', 'e', fr))
bp1 = bed.start1 if True else bed.end1
bp2 = bed.start2 if False else bed.end2
#choose orientation of the first read and automatically flip the second
orient1 = '5' if bed.orient1 == '-' else '3'
orient2 = '5' if bed.orient2 == '+' else '3'
j = [(bp1, orient1, bed.chrom1), (bp2, orient2, bed.chrom2)]; j.sort(); uBP1, dBP2 = j
uBreak, uDir, uRef = uBP1
dBreak, dDir, dRef = dBP2
j = [uRef, dRef]; j.sort(); refKey = "_".join(j)
is_reverse = bed.orient1 == bed.orient2
fr = Fr(False)
ret.append(FakeBread(bed.svtype, refKey, uRef, dRef, uBreak, dBreak, uDir, dDir, is_reverse, 'i', 'e', fr))
bp1 = bed.start1 if False else bed.end1
bp2 = bed.start2 if True else bed.end2
#choose orientation of the first read and automatically flip the second
orient1 = '5' if bed.orient1 == '-' else '3'
orient2 = '5' if bed.orient2 == '+' else '3'
j = [(bp1, orient1, bed.chrom1), (bp2, orient2, bed.chrom2)]; j.sort(); uBP1, dBP2 = j
uBreak, uDir, uRef = uBP1
dBreak, dDir, dRef = dBP2
j = [uRef, dRef]; j.sort(); refKey = "_".join(j)
is_reverse = bed.orient1 == bed.orient2
fr = Fr(False)
ret.append(FakeBread(bed.svtype, refKey, uRef, dRef, uBreak, dBreak, uDir, dDir, is_reverse, 'i', 'e', fr))
bp1 = bed.start1 if False else bed.end1
bp2 = bed.start2 if False else bed.end2
#choose orientation of the first read and automatically flip the second
orient1 = '5' if bed.orient1 == '-' else '3'
orient2 = '5' if bed.orient2 == '+' else '3'
j = [(bp1, orient1, bed.chrom1), (bp2, orient2, bed.chrom2)]; j.sort(); uBP1, dBP2 = j
uBreak, uDir, uRef = uBP1
dBreak, dDir, dRef = dBP2
j = [uRef, dRef]; j.sort(); refKey = "_".join(j)
is_reverse = bed.orient1 == bed.orient2
fr = Fr(False)
ret.append(FakeBread(bed.svtype, refKey, uRef, dRef, uBreak, dBreak, uDir, dDir, is_reverse, 'i', 'e', fr))
else:
#PE -> <-
#+/- to pick the breakpoint locations
bp1 = bed.start1 if bed.orient1 == '-' else bed.end1
bp2 = bed.start2 if bed.orient2 == '-' else bed.end2
#choose orientation of the first read and automatically flip the second
orient1 = '5' if bed.orient1 == '-' else '3'
orient2 = '5' if bed.orient2 == '+' else '3'
j = [(bp1, orient1, bed.chrom1), (bp2, orient2,bed.chrom2)]; j.sort(); uBP1, dBP2 = j
uBreak, uDir, uRef = uBP1
dBreak, dDir, dRef = dBP2
is_reverse = bed.orient1 == bed.orient2
fr = Fr(is_reverse)
#My main read I'll fake cluster with everything
ret.append(FakeBread(bed.svtype, refKey, uRef, dRef, uBreak, dBreak, uDir, dDir, is_reverse, 'i', 'e', fr))
#logging.debug(fakey)
return ret
def bedToFakey(bed):
"""
Make a fakeBread for bed entry
returns a list.
If bed.svtype == 'UNK',
then a fakey for every orientation
combination will be created
"""
ret = []
uRef = bed.chrom; dRef = bed.chrom
j = [uRef, dRef]; j.sort(); refKey = "_".join(j)
#PE -> <-
#+/- to pick the breakpoint locations
fr = Fr(False)
if bed.svtype == 'DEL' or bed.svtype == 'UNK':
ret.append(FakeBread(bed.svtype, refKey, uRef, dRef, bed.start, bed.end, '3', '3', False, 'i', 'e', fr))
elif bed.svtype == 'INS' or bed.svtype == 'UNK':
ret.append(FakeBread(bed.svtype, refKey, uRef, dRef, bed.start, bed.end, '3', '3', False, 'e', 'i', fr))
elif bed.svtype == 'MIS' or bed.svtype == 'UNK':
ret.append(FakeBread(bed.svtype, refKey, uRef, dRef, bed.start, bed.end, '3', '5', False, 'i', 'e', fr))
return ret
def removeRedundantReads(reads):
"""
check readname, chrom/pos, flag to see if you're looking at the same
reads
"""
ret = []
while len(reads) > 0:
cur = reads.pop()
rm = []
for i in range(len(reads)):
cmp = reads[i]
if cur.compare(cmp) == 0:
rm.append(cmp)
ret.append(cur)
for i in rm:
reads.remove(i)
return ret
#refactored
def tailsSearch(bam, fakey, args):
"""
Look in bam for reads that support the predicted structural variant
"""
reads = []
#First breakpoint -
fetchS = max(0, fakey.uBreak - args.fetchbuffer)
fetchE = min(fakey.uBreak + args.fetchbuffer, bam.lengths[bam.references.index(fakey.uRef)])
if fetchS > fetchE:#don't know why this happened
fetchS, fetchE = fetchE, fetchS
reads.extend([x for x in bam.fetch(fakey.uRef, fetchS, fetchE)])
#Second breakpoint -
fetchS = max(0, fakey.dBreak - args.fetchbuffer)
fetchE = min(fakey.dBreak + args.fetchbuffer, bam.lengths[bam.references.index(fakey.dRef)])
if fetchS > fetchE:#don't know why this happened
fetchS, fetchE = fetchE, fetchS
reads.extend([x for x in bam.fetch(fakey.dRef, fetchS, fetchE)])
#It's possible that the same reads are fetched from both breakpoints
#We need to remove those redundant reads
reads = removeRedundantReads(reads)
#This is the evidence we have
points, tlocs = tails.parseBreakReads(reads, getrname = bam.getrname)
#now we need to keep track of who is near our fakey
nears = []
for key in points:
for read in points[key]:
#search for the original
if read.near(fakey):
nears.append(read.toBriefString())
return len(reads) > 1, nears
def oldTailsSearch(bam, bed, args):
"""
Populate the answer dictionary by looking for tails
through the bam
Returns a list of pbsuite.honey.TGraf.Bnode that support
"""
fetchS = max(0, bed.start - args.fetchbuffer)
fetchE = min(bed.end + args.fetchbuffer, bam.lengths[bam.references.index(bed.chrom)])
points = tails.makeBreakReads(bam.fetch(bed.chrom, fetchS, fetchE), getrname = bam.getrname)
reads = []
anyCoverage = False
for key in points:
anyCoverage = True
#eventually will need tloc work
if key.split('_')[0] != bed.chrom:
continue
#eventually will need a reference allele check for tails
for read in points[key]:
anno = read.annotate()
if anno in ['TLOC', 'INV']:
anno = 'MIS'
#TLOCs...
if bed.chrom != bam.getrname(read.read.tid):
continue
if anno != bed.svtype:
#Not perfect..
continue
#within reciprocal ovl
maxStart = max(bed.start, read.uBreak)
minEnd = min(bed.end, read.dBreak)
if minEnd <= maxStart: #No overlap
continue
maxSpan = max(bed.end-bed.start, read.dBreak - read.uBreak)
recipOvl = abs(maxStart-minEnd) / float(maxSpan)
logging.debug("predictVar [%d:%d] - tailRead [%d:%d]" \
% (bed.start, bed.end, read.uBreak, read.dBreak))
if recipOvl < args.overlapbuffer:#not enough overlap
continue
anno = read.annotate()
reads.append(read.toBriefString())
#ret = ",".join(['t[%s]' % (str(x)) for x in reads])
return anyCoverage, reads
def spotsSearch_asm(bam, bed, args):
"""
find spots in high-accuracy contigs
I'm going to have a problem with Insertion offsets before the variant
if there are too many of them, I'm going to be effed
"""
#MIS types can't be resolved from spots
# EXCEPT, however, if they're actually INS/DEL
# except a single bp, they could
if bed.svtype == 'MIS':
return False, '?',[]
fetchS = max(0, bed.start - args.fetchbuffer)
fetchE = min(bed.end + args.fetchbuffer, bam.lengths[bam.references.index(bed.chrom)])
leeway = bed.size * args.sizebuffer
ref = '?'
vars = []
anyCoverage = False
for read in bam.fetch(bed.chrom, fetchS, fetchE):
anyCoverage = True
if read.pos > bed.start or read.aend < bed.end:
#Not spanning our region
logging.debug("%s doesn't span region" % (read.qname))
continue
ref = False
if bed.size + leeway > 50000:
logging.debug("Variant is too long (%dbp), we are assuming reference" % (read.qname, len(read.seq)))
continue
cigar = spots.expandCigar(read.cigar)
regionStart = max(read.pos, bed.start - args.maxDelta)
regionEnd = min(read.aend, bed.end + args.maxDelta)
readPosition = read.pos
c = "".join([str(x) for x in cigar])
logging.debug(c)
logging.debug(c[regionStart-read.pos : regionEnd-read.pos])
if bed.svtype == 'INS':
match = re.search("(^|[^1])1{%d,%d}([^1]|$)" % (bed.size-leeway, bed.size+leeway), c[regionStart-read.pos : regionEnd-read.pos])
elif bed.svtype == 'DEL':
match = re.search("(^|[^2])2{%d,%d}([^2]|$)" % (bed.size-leeway, bed.size+leeway), c[regionStart-read.pos : regionEnd-read.pos])
if match is None:
ref = True
else:
#subtract insertion errors to correct the offset
#Insertion offset subraction
subtract = cigar[:(regionStart-read.pos) + match.start()].count(1)
if bed.svtype == "INS":
pos = match.start() + read.pos + (regionStart - read.pos) - subtract
s,e = match.span(); size = e-s
var = Variant(bed.chrom, pos, pos + 1, bed.svtype, size)
if bed.svtype == "DEL":
subtract = cigar[:(regionStart-read.pos)+ match.start()].count(1)
spos = match.start() + read.pos + (regionStart -read.pos) - subtract
epos = match.end() + read.pos + (regionStart - read.pos) - subtract
s,e = match.span(); size = e-s
var = Variant(bed.chrom, spos, epos, bed.svtype, size)
vars.append(var)
if len(vars) > 0:
vars = str(vars[0]) + ("*%d" % len(vars))
else:
vars = ""
return anyCoverage, ref, vars
def spotsSearch(bam, bed, args):
"""
take a pysam.Samfile and fetch reads in chrom/start/end region
see if any reads support the call
But this doesn't take into account that I have specific groupIds to use...
"""
leeway = bed.size * args.sizebuffer
fetchS = max(0, bed.start - args.fetchbuffer)
fetchE = min(bed.end + args.fetchbuffer, bam.lengths[bam.references.index(bed.chrom)])
if fetchS > fetchE:
fetchS, fetchE = fetchE, fetchS
vars = []
ref = '?'
anyCoverage = False
for read in bam.fetch(bed.chrom, fetchS, fetchE):
if read.pos > bed.start or read.aend < bed.end:
#Not spanning our region
logging.debug("%s doesn't span region" % (read.qname))
continue
logging.debug("looking at %s" % (read.qname))
anyCoverage = True
ref = False #we now have the opportunity to find the reference
#I'm going to need md if I get good a MIS
regionStart = bed.start - args.maxDelta
regionEnd = bed.end + args.maxDelta
leeway = bed.size * args.sizebuffer
logging.debug(("regionStart, regionEnd, estSize, leeway, "
"estSize+leeway, estSize-leeway"))
logging.debug("%d %d %d %d %d %d" % (regionStart, regionEnd, \
bed.size, leeway, bed.size+leeway, bed.size-leeway))
foundVar = False
for svstart, svsize, svtype in spots.expandCigar(read, args.minErr, collapse=3, makeAlt=False):
if svstart >= regionStart and svstart <= regionEnd and \
bed.svtype == svtype and \
bed.size - leeway <= svsize <= bed.size + leeway:
#check overlap -- I like this logic
#if (var.start <= bed.start and bed.end <= var.end) \
#or (bed.start <= var.start and var.end <= bed.end) \
#or (bed.start <= var.end and var.end <= bed.end) \
#or (bed.start <= var.start and var.start <= bed.end):
#vars.append(var)
#foundVar = True
#else:
#maxS = max(var.start, bed.start)
#minE = min(var.end, bed.end)
#if abs(maxS-minE) <= args.maxDelta:
#vars.append(var)
#foundVar = True
foundVar = True
if svtype == "DEL":
vars.append(Variant(bed.chrom, svstart, svstart + svsize, svtype, svsize))
elif svtype == "INS":
vars.append(Variant(bed.chrom, svstart, svstart, svtype, svsize))
if not foundVar:#this might be broken
ref = True
#why only the first and not an average? HOMAlt... would suck
if len(vars) > 0:
vars = str(vars[0]) + ("*%d" % len(vars))
else:
vars = ""
return anyCoverage, ref, vars
#Here are some helper methods for parsing force annotation results
forceRe = re.compile("(?P<ref>True|False|\?)\[(?P<tails>.*)\|(?P<spots>.*)\]")
def parseForce(data):
"""
turns the force output into a dict
"""
if data == 'no_cov' or data == '.':
return None
#will fail on malformed entries
search = forceRe.search(data)
if search is not None:
d = search.groupdict()
else:
print "problem parsing", data
return 'prob'
if d["ref"] == '?':
d["ref"] = None
elif d["ref"] == 'True':
d["ref"] = True
elif d["ref"] == 'False':
d["ref"] = False
d["tails"] = [x for x in d["tails"].split(',') if x != '']
d["spots"] = [x for x in d["spots"].split(',') if x != '']
return d
def genoTyper(data):
"""
"""
if data["ref"] is not None:
if data["ref"]:
if len(data["tails"]) > 0 or len(data["spots"]) > 0:
genoType = "0/1"
else:
genoType = "0/0"
elif not data["ref"]:
if len(data["tails"]) > 0 or len(data["spots"]) > 0:
genoType = "1/1"
else:
genoType = "./."
else:
if len(data["tails"]) > 0 or len(data["spots"]) > 0:
genoType = "./1"
else:
genoType = "./."
return genoType
def run(args):
args = parseArgs(args)
bam = pysam.Samfile(args.bam)
#putative caller
fh = open(args.bed)
tails.BUFFER = args.maxDelta
#CTX and ITX are for breakpoints that have orientations
#UNK we'll try to find anything that matches (good debugging because we should only be finding
#support for one of the things we make most always)
numEntries = 0
for line in fh.readlines():
if line.startswith("#"):
continue
myentry = line.strip().split('\t')
if not args.bedPE:
myentry = BedEntry(*myentry)
else:
myentry = BedPEEntry(*myentry)
if myentry.svtype not in vtypes:
#if myentry.svtype == 'UNK':
#logging.warning("Bed Entry %s is UNK and can't be forced... skipping" % (repr(myentry)))
#sys.stdout.write(line.strip() + "\t.\n")
#continue
#else:
logging.error("Bed Entry %s svtype column isn't one of %s" % (repr(myentry), str(vtypes)))
exit(1)
if not args.bedPE:
if myentry.chrom not in bam.references:
logging.error("Invalid Chromosome %s" % myentry.chrom)
continue
mySupport = bedToFakey(myentry)
tailVars = []
anyCoverage1 = False
for i in mySupport:
ancov1, t = tailsSearch(bam, i, args)
anyCoverage1 = anyCoverage1 or ancov1
tailVars.extend(t)
if False:#args.asm:
anyCoverage2, foundRef, spotVars = spotsSearch_asm(bam, myentry, args)
else:
anyCoverage2, foundRef, spotVars = spotsSearch(bam, myentry, args)
else:
anyCoverage1 = False
tailVars = []
mySupport = bedpeToFakey(myentry)
for i in mySupport:
ancov1, t = tailsSearch(bam, i, args)
anyCoverage1 = anyCoverage1 or ancov1
tailVars.extend(t)
foundRef = False
anyCoverage2 = False
spotVars = ""
#Eventually, I can get rid of the True/False/? once the genotyper is finished
#I'm outputing the variant reads and if we found the ref (True, False, ?) where ? means no evidence for or
# against
if not anyCoverage1 and not anyCoverage2:
annot = "no_cov"
logging.info("no coverage")
else:
annot = "%s[%s|%s]" % (foundRef, ",".join(tailVars), spotVars)
#this is wrong. It's the string's length
logging.info("Found %d tailed, %d spotted reads" % (len(tailVars), len(spotVars)))
sys.stdout.write(line.strip() + "\t" + annot + "\n")
numEntries += 1
if numEntries % 250 == 0:
sys.stdout.flush()
if __name__ == '__main__':
run(sys.argv[1:])
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