"""complements the provided DNA sequence and returns it"""

global dna_comp

if dna_comp is None:

dna_comp = string.maketrans("ATCGMRWSYKNatcgmrwsykn",

"TAGCKYWSRMNtagckywsrmn")

if chrom not in genome_file.root:

known_chrom = [node.name for node in genome_file.root]

raise ValueError("unknown chromosome %s, possible chromosomes are: "

+ ",".join(known_chrom))

# numpy array representation of sequence and convert to string

chrom_node = genome_file.getNode("/%s" %chrom)

np_seq = chrom_node[start-1:end]

seq_str = str_from_nparray(np_seq)

if strand == "-":

# reverse-complement sequence

seq_str = revcomp(seq_str)

guideRna = []

#has list with RNAs with proper direction

guideRnaOutput = []

location = []

direction = []

i = 0

while i < (len(seq)-1):

if seq[i] == "G":

#print "HI"

if seq[i+1] == "G":

#print i

#only if we are at least 23 bases in can we store a guide RNA, so check and store in list

if i >= gRNA_length+1:

if seq[i-(gRNA_length+1)] == "G":

guideRna.append(seq[i-(gRNA_length+1):i-1])

guideRnaOutput.append(seq[i-(gRNA_length+1):i-1])

#guideRnaPam.append(seq[i-21:i-1] + " " + seq[i-1:i+2])

start = (i - (gRNA_length+1)) + inStart

end = (i - 1) + inStart

location.append(str(start) + '-' + str(end))

direction.append('fwd')

#increment counter

i = i + 1

## add loop to look for CCN start once found take reverse complement

w = 0

forward = ""

while w < (len(seq)-1):

if seq[w] == "C":

if seq[w+1] == "C":

#print w

#print len(seq)

#only if we are at least 23 bases in can we store a guide RNA, so check and store in list

if (w+(gRNA_length+3)) <= (len(seq)):

#add check here for N

forward = seq[w+3: w+(gRNA_length+3)]

#print forward

CODE={'A':'T','T':'A','C':'G','G':'C','N':'N'}

minus_seq=''

for c in forward:

minus_seq=minus_seq+CODE[c]

reverse_seq=minus_seq[::-1]

#print reverse_seq

if reverse_seq[:1] == "G":

guideRnaOutput.append(reverse_seq)

guideRna.append(forward)

#guideRnaPam.append(seq[w:w+3] + " " + seq[w+3:w+23])

start = w+3+inStart

end = w+(gRNA_length+3)+inStart

location.append(str(start) + '-' + str(end))

direction.append('rev')

#increment counter

w = w + 1

#print guideRna

#print guideRnaPam

#print guideRnaOutput

#print location

#print direction

b = 0

while b < len(guideRna):

if "N" in guideRna[b]:

del guideRna[b]

del guideRnaOutput[b]

del location[b]

del direction[b]

b = b + 1

else:

b = b +1

#write to file

print len(guideRna)

if len(sys.argv) < 2:

sys.stderr.write("usage: %s <chrom> [<start> <end>]\n" % sys.argv[0])

exit(2)

inChrom = sys.argv[1]

if len(sys.argv) > 2:

inStart = int(sys.argv[2])

inEnd = int(sys.argv[3])

else:

start = ""

end = ""

sys.stderr.write("%s %d %d\n" % (inChrom, inStart, inEnd))

inStrand = "+"

#rename depending on input gene

fastainput = open("Input_GATA3.fasta", 'w')

#change path to hdf5 file you want to use

seq_h5 = tables.openFile("/iblm/netapp/data1/external/GRC37/GRC37.h5", "r")

inputRegion = get_seq(inChrom, inStart, inEnd, inStrand, seq_h5)

#print inputRegion

fastainput.write(">" + inChrom + '\n')

fastainput.write(inputRegion)

fastainput.close()

#reopen the fasta input file from above

f = open("Input_GATA3.fasta", "r")

#declare chr start and end for region that you want to extract guides from

file = f.readlines()

#Sguide = []

#guideList = []

#create output file

#rename depending on gene of interest

outfile = open('GuideRNAs_GATA3','w')

outfile2 = open('FQGuides_GATA3.fq' , 'w')

#can change seq and header to lists if there are more than one header in fasta file with several sequences

#declare empty lists

sequence = []

header = []

seq = ""

header = ""

GC = []

#store all the headers in a list

for f in file:

if f.startswith('>'):

header = header + f

header = header.splitlines()[0]

#get ride of new line charaters and spaces

else:

f = f.replace(" ", "")

f = f.replace("\n", "")

seq = seq + f

#i = 0

#make it all upper case, easier to parse

seq = seq.upper()

#print guideRna

#call function to find all guide RNAs

gRNA_length = 20

guideRna1, guideRnaOutput1, location1, direction1 = finding_guides(seq,inStart, inEnd,gRNA_length)

gRNA_length = 19

guideRna2,guideRnaOutput2, location2, direction2 = finding_guides(seq,inStart, inEnd,gRNA_length)

gRNA_length = 21

guideRna3, guideRnaOutput3, location3, direction3 = finding_guides(seq,inStart, inEnd,gRNA_length)

guideRna = guideRna1 + guideRna2 + guideRna3

guideRnaOutput = guideRnaOutput1 + guideRnaOutput2 + guideRnaOutput3

location = location1 + location2 + location3

direction = direction1 + direction2 + direction3

print guideRna

print guideRnaOutput

print len(guideRna)

#create .fq file used for alignment

j = 0

outfile.write(header)

while j<len(guideRna):

outfile.write(guideRna[j] + '\n')

outfile2.write('@' + str(j) + '\n')

outfile2.write(guideRna[j] + '\n')

outfile2.write('+' + '\n')

score = "I"*len(guideRna[j])

outfile2.write(score + '\n')

j = j+1

#if we don't close and reopen FQGuides.fq we cannot excute the commands since FQGuides.fq was only a write file

outfile2.close()

#reopen the .fq file created above

open("FQGuides_GATA3.fq", "r")

print "We created a .fq file with all the guides"

#call bwa aligner from this code and get information from the sam file

#bottleneck here

#change input reference genome and output file name here

cmd = 'bwa aln -o 0 -n 3 -N -t 5 hg37.fa FQGuides_GATA3.fq > aln_GATA3.sai'

os.system(cmd)

print "Alignment Step has finished"

#change input reference genome and output file name here

cmd2 = 'bwa samse -n 1000 hg37.fa aln_GATA3.sai FQGuides_GATA3.fq > aln_GATA3.sam'

os.system(cmd2)

print "Sam file has been created"

import re