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ssCorrect.py
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ssCorrect.py
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from __future__ import print_function
########################################################################
# File: ssCorrect.py
# executable: ssCorrect.py
# Purpose:
#
#
# Author: Cameron M. Soulette
# History: cms 05/01/2018 Created
#
########################################################################
########################################################################
# Hot Imports & Global Variable
########################################################################
import os, sys
import numpy as np
import multiprocessing
from multiprocessing import Pool
from contextlib import closing
from intervaltree import Interval, IntervalTree
import random
from tqdm import *
########################################################################
# CommandLine
########################################################################
class CommandLine(object) :
'''
Handle the command line, usage and help requests.
CommandLine uses argparse, now standard in 2.7 and beyond.
it implements a standard command line argument parser with various argument options,
and a standard usage and help,
attributes:
myCommandLine.args is a dictionary which includes each of the available command line arguments as
myCommandLine.args['option']
methods:
'''
def __init__(self, inOpts=None) :
'''
CommandLine constructor.
Implements a parser to interpret the command line argv string using argparse.
'''
import argparse
self.parser = argparse.ArgumentParser(description = ' ssCorrect.py - a tool to leverage annotation and short read data to correct misaligned splice junctions in short read data.',
epilog = 'Please feel free to forward any questions/concerns to /dev/null',
add_help = True, #default is True
prefix_chars = '-',
usage = '%(prog)s -i reads.bed -g annotations.gtf -j other_junctions.bed -o out_file.bed')
# Add args
self.parser.add_argument('-i', "--input_bed", action = 'store', required=True, help='Input reads in bed12 format.')
self.parser.add_argument('-g', "--gtf", action = 'store', required=False, help='Gencode annotation file.')
self.parser.add_argument('-j', "--junctionsBed", default=None, action = 'store', required=False, help='Short-read supported junctions in bed6 format (Optiona) [BED entries must be UNIQUE and have strand information].')
self.parser.add_argument('-w', '--wiggleWindow', action = 'store', type=int, required=False, default = 15, help='Splice site correction window flank size.')
self.parser.add_argument('-o', "--output_fname", action = 'store', required=True, help='Output file name.')
# (Under Development!!!)
#self.parser.add_argument('--report_junctions', action = 'store', required=False, default= "norm", choices=['strict', 'norm', 'all'],
# help = """Choose which types of nanopore splice sites to report:
# "strict" - SS must be in gtf or junctionsBed, and have a single closest hit.
# "norm" - SS must be in gtf or junctionsBed (tie for closest hit is random).
# "all" - SS can be in gtf, junctionsBed, or unique to nanopore data.""")
self.parser.add_argument('-p', "--threads", action = 'store', required=False, type=int, default = 2, help='Number of threads.')
self.parser.add_argument('--keepZero', action = 'store_true', required=False, default = False, help='Keep alignments with no spliced junctions (single exon txns).')
self.parser.add_argument("--quiet", action = 'store_false', required=False, default = True, help='Do not display progress')
self.parser.add_argument("--cleanup", action = 'store_true', required=False, default = False, help='Remove teomprary files with correction info.')
if inOpts is None :
self.args = vars(self.parser.parse_args())
else :
self.args = vars(self.parser.parse_args(inOpts))
########################################################################
# BED File
########################################################################
class BED12(object):
'''
Handles BED format file input and output.
# BED12 has some built in functions
# for formally defining elemnts of a bed12
# and to preform coordinate conversions
Attribute names are stable, but the value refreshes while iterating through
the bed file.
attributes:
chrom, start, end, strand = reference aligment descriptors
read = query/feature id
score = integer
c1, c2 = (formally) describe where the open reading frame starts and stop
exons, size, starts = bed12 alignment blocks descibing where the aligmnents matches in the reference.
methods:
getLine - gets line from bed file, and defines values for attributes
bed12to(Juncs|Exons) - converts bed12 aligmnet blocks to reference coordinate positions
getLine must be called before bed12to(Juncs|Exons) can be called since it relies on attributes defined in getLine.
'''
def __init__(self, fname=None):
self.fname = fname
if not os.path.isfile(fname):
print("%s does not exist. Exiting.", file=sys.stderr)
sys.exit(1)
def getLine(self):
with open(self.fname,'r') as entries:
for entry in entries:
cols = entry.rstrip().split()
self.chrom, self.start, self.end, self.name = cols[0], int(cols[1]), int(cols[2]), cols[3]
self.score, self.strand, self.c1, self.c2 = int(cols[4]), cols[5], int(cols[6]), int(cols[7])
self.color, self.exons = cols[8], int(cols[9]),
self.sizes = [int(x) for x in cols[10].split(",")[:-1]] if cols[10][-1] == "," else [int(x) for x in (cols[10]+",").split(",")[:-1]]
self.starts = [int(x) for x in cols[11].split(",")[:-1]] if cols[11][-1] == "," else [int(x) for x in (cols[11]+",").split(",")[:-1]]
yield cols
def bed12toJuncs(self):
'''
Take bed12 entry and convert block/sizes to junction coordinates.
'''
junctions = list()
for num, st in enumerate(self.starts,0):
if num+1 >= len(self.starts):
break
ss1 = self.start + st + self.sizes[num]
ss2 = self.start + self.starts[num+1]
junctions.append((ss1,ss2))
return junctions
def bed12toExons(self):
'''
Take bed12 entry and convert block/sizes to exon coordinates.
'''
exons = list()
for num, st in enumerate(self.starts,0):
c1 = self.start + st
c2 = c1 + self.sizes[num]
exons.append((c1,c2))
return exons
def determinType(self):
pass
########################################################################
# Functions
########################################################################
def juncsToBed12(start, end, coords):
'''
Take alignment start, end, and junction coords and convert to block/size bed12 format.
start = integer
end = integer
coords = list formatted like so [(j1_left,j1_right),(j2_left,j2_right)]
'''
novelFlag = False
sizes, starts = [],[]
# initial start is 0
if len(coords) > 0:
for num,junc in enumerate(coords,0):
ss1, ss2, novel1, novel2 = junc
if novel1 or novel2: novelFlag = True
if num == 0:
st = 0
size = abs(start-ss1)
else:
st = coords[num-1][1] - start
size = ss1 - (st + start)
starts.append(st)
sizes.append(size)
st = coords[-1][1] - start
size = end - (st + start)
starts.append(st)
sizes.append(size)
return len(starts), sizes, starts, novelFlag
else:
return 1, [end-start], [0], novelFlag
def buildOtherDB(acceptors, donors, spliceSites, bedJuncs, wiggle, proc):
lineNum = 0
with open(bedJuncs,'r') as bedLines:
lineNum +=1
cols = next(bedLines).split()
for i in bedLines:
lineNum += 1
# guess what kind of bedFile
if cols[-1] == "+" or cols[-1] == "-":
# normal bed
reverseSS = "-"
strandCol = -1
starOffset = 0
elif len(cols) == 12:
# bed12
bedType = "bed12"
print("ERROR: Bed12 not currently supported for other_juncs.bed. Please convert to bed6. Exiting.", file=sys.stderr)
elif cols[3] == "0" or cols[3] == "1" or cols[3] == "2":
# star junc.tab
reverseSS = "2"
strandCol = 3
starOffset = 1
else:
print("ERROR: Cannot find strand info for %s. Is this bed6 or STAR_juncs.tab file? Exit." % bedJuncs, file=sys.stderr)
proc += 1
with open(bedJuncs,'r') as bedLines:
for line in tqdm(bedLines, total=lineNum, desc="Adding splice sites from %s to interval tree" % bedJuncs, dynamic_ncols=True, position = proc) if verbose else bedLines:
cols = line.rstrip().split()
chrom, c1, c2, strand = cols[0], int(cols[1])-starOffset, int(cols[2]), cols[strandCol]
if strand == reverseSS: c1,c2 = c2,c1
if chrom not in donors:
donors[chrom] = IntervalTree()
acceptors[chrom] = IntervalTree()
if (chrom,c1) not in spliceSites:
donors[chrom][c1-wiggle:c1+wiggle] = ('other',c1, strand)
if (chrom,c2) not in spliceSites:
acceptors[chrom][c2-wiggle:c2+wiggle] = ('other',c2, strand)
return donors, acceptors
def buildGTFDB(file, wiggle, proc):
exons = dict()
junctionSet = set()
if verbose: print("reading gtf %s ..." % (file), file=sys.stderr)
with open(file,'r') as lines:
for l in lines:
if l[0] == "#": # skip header lines
continue
cols = l.split("\t")
if "exon" == cols[2]:
# -1 for 1 to 0 based conversion
chrom, c1, c2, strand = cols[0], int(cols[3])-1, int(cols[4]), cols[6]
#txn info is in the SECOND position of the shoutout column
txn = cols[-1].split(";")[1].split()[-1].replace('"','')
key = (chrom, txn, strand)
if key not in exons:
exons[key] = list()
exons[key].append(c1)
exons[key].append(c2)
txnList = list(exons.keys())
proc += 1
for exonInfo in tqdm(txnList, total=len(txnList), desc="Building junction interval tree from GTF", dynamic_ncols=True, position=proc) if verbose else txnList:
chrom, txn, strand = exonInfo
coords = exons[exonInfo]
# assume lowest and highest as TSS and TES, and remove them
coords.sort()
coords = coords[1:-1]
if len(coords)<2: continue
if chrom not in donors: #make the interval tree
donors[chrom] = IntervalTree()
acceptors[chrom] = IntervalTree()
for pos in range(0,len(coords)-1,2):
c1 = coords[pos]
c2 = coords[pos+1]
if strand == "-": c1,c2 = c2,c1
if (chrom,c1) not in junctionSet:
donors[chrom][c1-wiggle:c1+wiggle] = ('gtf',c1,strand)
junctionSet.add((chrom,c1))
if (chrom,c2) not in junctionSet:
acceptors[chrom][c2-wiggle:c2+wiggle] = ('gtf',c2,strand)
junctionSet.add((chrom,c2))
return donors, acceptors, junctionSet
def resolveHits(spliceSite, hits, read):
hits = list(hits)
if read == "30e93c3f-28c2-4b3":
print(hits)
print(spliceSite)
if len(hits)<1:
# novel
return (spliceSite, spliceSite, np.nan, np.nan, True)
elif len(hits) == 1:
# single closest hit
return (hits[0].data[1],spliceSite,spliceSite-hits[0].data[1], hits[0].data[0], False)
else:
# multi hit
distances = [(abs(spliceSite-hit.data[1]),hit.data[1],hit.data[0]) for hit in hits]
sortedDist = sorted(distances,key=lambda x: x[0])
top, second = sortedDist[0], sortedDist[1]
if top[0] == second[0]:
best = random.choice([top,second])
distance, ssCord, refType = best
else:
distance, ssCord, refType = top
return (ssCord,spliceSite,spliceSite-ssCord,refType, False)
def ssCorrect(chrom, bedFile, fileSize, procNum):
data = BED12(bedFile)
statsOut = open("%s_ssCorrectionInfo.tsv" % chrom,'w')
tempOut = open("%s_corrected.bed" % chrom ,'w')
tempNovelOut = open("%s_uncorrected.bed" % chrom ,'w')
for line in tqdm(data.getLine(), total=fileSize, desc="Working on %s" % chrom, position=procNum,dynamic_ncols=True) if verbose else data.getLine():
junctionCoords = data.bed12toJuncs()
ch, st, end, blocks, strand = data.chrom, data.start, data.end, data.exons, data.strand
readID = data.name[:18]
if ch in donors and ch in acceptors:
if strand == "-":
hits = [ (resolveHits(x[0],donors[ch][x[0]],readID), resolveHits(x[1], acceptors[ch][x[1]],readID))
for x in junctionCoords]
else:
hits = [ (resolveHits(x[0],acceptors[ch][x[0]],readID), resolveHits(x[1], donors[ch][x[1]],readID))
for x in junctionCoords]
correctedJuncs = [(x[0][0],x[1][0],x[0][-1],x[1][-1]) for x in hits]
else:
# un annotated reference chrom/contig
if len(junctionCoords) < 1:
if keepZero:
print(data.chrom, data.start, data.end, readID,
data.score, data.strand, data.c1, data.c2, data.color,
data.exons, "%s," % data.sizes[0], "%s," % data.starts[0], sep="\t",file=tempNovelOut)
else:
print(data.chrom, data.start, data.end, readID,
data.score, data.strand, data.c1, data.c2, data.color,
data.exons, "%s," % data.sizes[0], "%s," % data.starts[0], sep="\t",file=tempNovelOut)
continue
if len(junctionCoords) < 1:
if keepZero:
print(data.chrom, data.start, data.end, readID,
data.score, data.strand, data.c1, data.c2, data.color,
data.exons, "%s," % data.sizes[0], "%s," % data.starts[0], sep="\t",file=tempOut)
continue
else:
exons, sizes, starts, novelJuncs = juncsToBed12(data.start,data.end,correctedJuncs)
if novelJuncs:
print(data.chrom, data.start, data.end, readID,
data.score, data.strand, data.c1, data.c2, data.color,
exons, ",".join(map(str,sizes))+",", ",".join(map(str,starts))+",", sep="\t",file=tempNovelOut)
else:
print(data.chrom, data.start, data.end, readID,
data.score, data.strand, data.c1, data.c2, data.color,
exons, ",".join(map(str,sizes))+",", ",".join(map(str,starts))+",", sep="\t",file=tempOut)
for i in hits:
left,right = i
if data.strand == "+":
print(readID, ch, "\t".join(map(str,left)), "5'", data.strand, sep="\t", file=statsOut)
print(readID, ch, "\t".join(map(str,right)), "3'", data.strand, sep="\t", file=statsOut)
else:
print(readID, ch, "\t".join(map(str,left)), "3'",data.strand, sep="\t", file=statsOut)
print(readID, ch, "\t".join(map(str,right)), "5'",data.strand, sep="\t", file=statsOut)
os.remove("%s_reads.temp.bed" % chrom)
statsOut.close()
tempOut.close()
tempNovelOut.close()
return ("%s_ssCorrectionInfo.tsv" % chrom, "%s_corrected.bed" % chrom, "%s_uncorrected.bed" % chrom)
def runCMD(x):
chrom, bedFile, fileSize, procNum = x
return ssCorrect(chrom, bedFile, fileSize,procNum)
def main():
'''
maine
'''
# Command Line Stuff...
myCommandLine = CommandLine()
bed = myCommandLine.args['input_bed']
gtf = myCommandLine.args['gtf']
otherJuncs = myCommandLine.args['junctionsBed']
wiggle = myCommandLine.args['wiggleWindow']
threads = myCommandLine.args['threads']
out = myCommandLine.args['output_fname']
cleanup = myCommandLine.args['cleanup']
# keep track of process number
proc = -1
global keepZero
keepZero = myCommandLine.args['keepZero']
#report = myCommandLine.args['report_junctions']
# There are a few functions that evaluate what verbose is defined as.
# Instead of passing it around, just global it.
global verbose
verbose = myCommandLine.args['quiet']
# Build interval tree with user splice site input data.
global donors, acceptors
donors, acceptors = dict(), dict()
donors, acceptors, spliceSites = buildGTFDB(gtf, wiggle, proc)
if otherJuncs != None: donors, acceptors = buildOtherDB(donors, acceptors, spliceSites, otherJuncs, wiggle, proc)
# Split bed by CHROMOSOME!!!!!
# and Check how large bed file to be correctd is.
fileDict = dict()
fileSizeDict = dict()
proc += 1
with open(bed) as f:
for l in tqdm(f, desc="Splitting isoforms by chrom for multiprocessing", position=proc,dynamic_ncols=True) if verbose else f:
chrom = l.split()[0]
if chrom not in fileDict:
fileDict[chrom] = open("%s_reads.temp.bed" % chrom,'w')
fileSizeDict[chrom] = 0
print(l.rstrip(), file=fileDict[chrom])
fileSizeDict[chrom] += 1
cmdList = list()
files = sorted(list(fileDict.keys()))
for key in files:
proc += 1
cmdList.append((key, "%s_reads.temp.bed" % key, fileSizeDict[key], proc))
fileDict[key].close()
# BED12 has some built in functions
# for formally defining elemnts of a bed12
# and to preform coordinate conversions
finalFiles = list()
proc += 1
with closing(Pool(threads)) as p:
for i in tqdm(p.imap(runCMD, cmdList), total=len(cmdList), desc="Correcting junctions per chromosome", position=proc,dynamic_ncols=True) if verbose else p.imap(runCMD, cmdList):
finalFiles.append(i)
proc += 1
allUncorrected = open("uncorrected_%s" % out, 'w')
with open(out,'w') as outF:
for i in tqdm(finalFiles, total=len(finalFiles), desc="Writing corrected juncs to %s" % out, dynamic_ncols=True, position = proc) if verbose else finalFiles:
stats,corrected, uncorrected = i
with open(corrected) as lines:
[print(x.rstrip(), file=outF) for x in lines]
with open(uncorrected) as lines:
[print(x.rstrip(), file=allUncorrected) for x in lines]
os.remove(uncorrected)
os.remove(corrected)
if cleanup:
os.remove(stats)
if __name__ == "__main__":
main()