def homologous_recombination_proposal(self, cutsite, donor_sequence):
cutsite = cutsite - 1
# check orientation and verify at least 15nt matching on both ends of ODN
if reverse_complement(donor_sequence[:self.__class__.MIN_HOMOLOGY_ARM]
) in self.wt_basecalls:
donor_sequence = reverse_complement(donor_sequence)
if donor_sequence[:self.__class__.MIN_HOMOLOGY_ARM] in self.wt_basecalls and \
donor_sequence[-self.__class__.MIN_HOMOLOGY_ARM:] in self.wt_basecalls:
# todo confirm that cutsite is inside the HDR region
donor_alignment = PairAlignment(self.wt_basecalls, donor_sequence)
donor_alignment.align_ssodn()
proposal_bases, proposal_trace, changed_bases, odn_start_pos = self.recombined_outcome_sequence(
donor_alignment.all_aligned_seqs, donor_sequence)
ep = EditProposal()
ep.sequence_data = proposal_bases
ep.trace_data = proposal_trace
ep.cutsite = cutsite
ep.summary = ProposalBase.HDR
ep.bases_changed = ProposalBase.HDR
return ep, changed_bases, odn_start_pos, donor_alignment
else:
raise Exception(
"Homology arms of length {} not found in control sequence".
format(self.__class__.MIN_HOMOLOGY_ARM))
def single_cut_edit_proposal(self,
cutsite,
label,
del_before=0,
del_after=0,
insertion=0):
cutsite = cutsite - 1
proposal_bases = []
proposal_trace = []
# deletion case
if del_before > 0 or del_after > 0:
deleted_bases = [cutsite - i for i in range(del_before)
] + [cutsite + i + 1 for i in range(del_after)]
for idx, base in enumerate(self.wt_basecalls):
if idx in deleted_bases:
proposal_base = ProposalBase('-', ProposalBase.DELETION,
idx)
else:
proposal_base = ProposalBase(base, ProposalBase.WILD_TYPE,
idx)
for base_index, base_color in enumerate(self.base_order):
proposal_trace.append(self.wt_trace[base_color][idx])
proposal_bases.append(proposal_base)
ep = EditProposal()
ep.sequence_data = proposal_bases
ep.cutsite = cutsite
ep.bases_changed = -(del_before + del_after)
ep.summary = '{}[{}]'.format(-(del_before + del_after), label)
ep.summary_json = {
'total': ep.bases_changed,
'details': [{
'label': label,
'value': ep.bases_changed
}]
}
ep.trace_data = proposal_trace
# insertion case
elif insertion > 0:
for idx, base in enumerate(self.wt_basecalls):
if idx == cutsite:
proposal_bases.append(
ProposalBase(base, ProposalBase.WILD_TYPE, idx))
for base_index, base_color in enumerate(self.base_order):
proposal_trace.append(self.wt_trace[base_color][idx])
for i in range(insertion):
proposal_bases.append(
ProposalBase('n', ProposalBase.INSERTION, idx))
for base_index, base in enumerate(self.base_order):
proposal_trace.append(0.25)
else:
proposal_bases.append(
ProposalBase(base, ProposalBase.WILD_TYPE, idx))
for base_index, base_color in enumerate(self.base_order):
proposal_trace.append(self.wt_trace[base_color][idx])
ep = EditProposal()
ep.sequence_data = proposal_bases
ep.cutsite = cutsite
ep.bases_changed = insertion
ep.summary = '{}[{}]'.format(insertion, label)
ep.summary_json = {
'total': ep.bases_changed,
'details': [{
'label': label,
'value': ep.bases_changed
}]
}
ep.trace_data = proposal_trace
# wildtype case
else:
for idx, base in enumerate(self.wt_basecalls):
proposal_base = ProposalBase(base, ProposalBase.WILD_TYPE, idx)
proposal_bases.append(proposal_base)
for base_index, base_color in enumerate(self.base_order):
proposal_trace.append(self.wt_trace[base_color][idx])
ep = EditProposal(wildtype=True)
ep.sequence_data = proposal_bases
ep.cutsite = cutsite
ep.bases_changed = 0
ep.summary = '0[{}]'.format(label)
ep.summary_json = {'total': ep.bases_changed, 'details': []}
ep.trace_data = proposal_trace
return ep
def multiplex_proposal(self,
cutsite1,
cutsite2,
label1,
label2,
cut1_del=(0, 0),
cut1_ins=0,
cut2_del=(0, 0),
cut2_ins=0,
dropout=False):
"""
Makes proposals for two guides cutting at the same time. This method handles both two separate indel proposals
or a dropout proposals. Both sites must be part of the edit and cutsite1 must come before cutsite2.
:param cutsite1: position of first cutsite
:param cutsite2: position of second cutsite
:param label1: label of guide target that creates first cutsite
:param label2: label of guide target that creates second cutsite
:param cut1_del: deletion size before and after first cutsite
:param cut1_ins: insertion size at first cutsite
:param cut2_del: deletion size before and after second cutsite
:param cut2_ins: insertion size at first cutsite
:param dropout: if the proposal is a dropout (removal of all bases between cutsites)
:return: EditProposal instance
"""
if cutsite2 <= cutsite1:
raise Exception(
'cutsite1 must come before cutsite2 values are ({}, {})'.
format(cutsite1, cutsite2))
cutsite1 -= 1
cutsite2 -= 1
proposal_bases = []
proposal_trace = []
summary_code = 'm' # multiplex
deleted_bases = []
if dropout:
summary_code = 'md' # multiplex dropout
for i in range(cutsite1 + 1, cutsite2 + 1):
deleted_bases.append(i)
# both cutsites results in deletions
# TODO, there are no safeguards for if the deletions for one cutsite exceed the boundaries of the other cutsite
# that info matters a lot for the summary
# for cut1 allow all deletions before
# for cut2 allow all deletions after
# for cut1 deletions after, stop if cut2 reached
# for cut2 deletions before, stop if cut1 reached
if cut1_del != (0, 0) and cut2_del != (0, 0):
# TODO, this should also handle the straight dropout case w/ no extra deletions
if dropout:
# zero out deletions between cutsites because dropout logic will take these deleted bases into account
# without double counting deletions
cut1_del_after = 0
cut2_del_before = 0
else:
cut1_del_after = cut1_del[1]
cut2_del_before = cut2_del[0]
cut1 = (cutsite1, cut1_del[0], cut1_del_after)
cut2 = (cutsite2, cut2_del_before, cut2_del[1])
for cutsite, del_before, del_after in [cut1, cut2]:
deleted_bases += [cutsite - i for i in range(del_before)] + [
cutsite + i + 1 for i in range(del_after)
]
for idx, base in enumerate(self.wt_basecalls):
if idx in deleted_bases:
proposal_base = ProposalBase('-', ProposalBase.DELETION,
idx)
else:
proposal_base = ProposalBase(base, ProposalBase.WILD_TYPE,
idx)
for base_index, base_color in enumerate(self.base_order):
proposal_trace.append(self.wt_trace[base_color][idx])
proposal_bases.append(proposal_base)
ep = EditProposal()
ep.sequence_data = proposal_bases
ep.cutsite = cutsite1
ep.cutsite2 = cutsite2
total_deleted = -cut1_del[
0] - cut1_del_after - cut2_del_before - cut2_del[1]
if dropout:
total_deleted += -(cutsite2 - cutsite1)
ep.bases_changed = total_deleted
cut1_del_size = cut1_del[0] + cut1_del_after
cut2_del_size = cut2_del_before + cut2_del[1]
ep.summary = '{}:{}-{}[{}],-{}[{}]'.format(total_deleted,
summary_code,
cut1_del_size, label1,
cut2_del_size, label2)
summary_json = {}
summary_json['total'] = ep.bases_changed
summary_json['details'] = []
if cut1_del_size > 0:
summary_json['details'].append({
'label': label1,
'value': -cut1_del_size
})
if cut2_del_size > 0:
summary_json['details'].append({
'label': label2,
'value': -cut2_del_size
})
if dropout:
summary_json['details'].append({
'label': 'dropout',
'value': cutsite1 - cutsite2
})
ep.summary_json = summary_json
ep.trace_data = proposal_trace
# insertion case
elif cut1_ins != 0 or cut2_ins != 0:
if dropout:
cut2_ins = 0
cut1 = (cutsite1, cut1_ins)
cut2 = (cutsite2, cut2_ins)
for idx, base in enumerate(self.wt_basecalls):
if idx in deleted_bases:
proposal_bases.append(
ProposalBase('-', ProposalBase.DELETION, idx))
# if base is in range where we need to do insertion
elif idx in [cutsite1, cutsite2]:
for cutsite, insertion_length in [cut1, cut2]:
if cutsite == idx:
proposal_bases.append(
ProposalBase(base, ProposalBase.WILD_TYPE,
idx))
for base_index, base_color in enumerate(
self.base_order):
proposal_trace.append(
self.wt_trace[base_color][idx])
for i in range(insertion_length):
proposal_bases.append(
ProposalBase('n', ProposalBase.INSERTION,
idx))
for base_index, base in enumerate(
self.base_order):
proposal_trace.append(0.25)
else:
proposal_bases.append(
ProposalBase(base, ProposalBase.WILD_TYPE, idx))
for base_index, base_color in enumerate(self.base_order):
proposal_trace.append(self.wt_trace[base_color][idx])
ep = EditProposal()
ep.sequence_data = proposal_bases
ep.cutsite = cutsite1 + cut1_ins
ep.cutsite2 = cutsite1 + cut1_ins + (cutsite2 -
cutsite1) + cut2_ins
ep.bases_changed = cut1_ins + cut2_ins
if dropout:
ep.bases_changed -= (cutsite2 - cutsite1)
ep.summary = '{}:{}+{}[{}],+{}[{}]'.format(ep.bases_changed,
summary_code, cut1_ins,
label1, cut2_ins,
label2)
summary_json = {}
summary_json['total'] = ep.bases_changed
summary_json['details'] = []
if cut1_ins > 0:
summary_json['details'].append({
'label': label1,
'value': cut1_ins
})
if cut2_ins > 0:
summary_json['details'].append({
'label': label2,
'value': cut2_ins
})
if dropout:
summary_json['details'].append({
'label': 'dropout',
'value': cutsite1 - cutsite2
})
ep.summary_json = summary_json
ep.trace_data = proposal_trace
ep.trace_data = proposal_trace
# the intervening sequence is dropped out and no bases inserted or deleted
else:
if dropout:
for idx, base in enumerate(self.wt_basecalls):
if idx in deleted_bases:
proposal_base = ProposalBase('-',
ProposalBase.DELETION,
idx)
else:
proposal_base = ProposalBase(base,
ProposalBase.WILD_TYPE,
idx)
for base_index, base_color in enumerate(
self.base_order):
proposal_trace.append(
self.wt_trace[base_color][idx])
proposal_bases.append(proposal_base)
ep = EditProposal()
ep.sequence_data = proposal_bases
ep.cutsite = cutsite1
ep.cutsite2 = cutsite2
total_deleted = -(cutsite2 - cutsite1)
ep.bases_changed = total_deleted
ep.summary = '{}:{}-0[{}],-0[{}]'.format(
total_deleted, summary_code, label1, label2)
ep.bases_changed = total_deleted
ep.summary_json = {
'total': ep.bases_changed,
'details': [{
'label': 'dropout',
'value': total_deleted
}]
}
ep.trace_data = proposal_trace
else:
# wild type case, use the single edit to model this case
return None
return ep
def multiplex_proposal(self,
cutsite1,
cutsite2,
label1,
label2,
cut1_del=(0, 0),
cut1_ins=0,
cut2_del=(0, 0),
cut2_ins=0,
dropout=False):
'''
this method will be used for two guides cutting at the same time
Note that both sites MUST be edited with either a deletion or an insertion.
Cases where one guide did not result in an edit should be handled by the single edit method
:param cutsite1: must come before cutsite2
:param cutsite2:
:param cut1_del:
:param cut1_ins:
:param cut2_del:
:param cut2_ins:
:return:
'''
cutsite1 = cutsite1 - 1
cutsite2 = cutsite2 - 1
proposal_bases = []
proposal_trace = []
if cutsite2 <= cutsite1:
raise Exception(
"cutsite1 must come before cutsite2 values are ({}, {})".
format(cutsite1, cutsite2))
summary_code = "m"
deleted_bases = []
if dropout:
summary_code = "md"
for i in range(cutsite1 + 1, cutsite2 + 1):
deleted_bases.append(i)
# deleted_bases = [cutsite-i for i in range(del_before)] + [cutsite+i+1 for i in range(del_after)]
# both cutsites results in deletions
# todo, there are no safeguards for if the deletions for one cutsite exceed the boundaries of the other cutsite
# that info matters a lot for the summary
# for cut1 allow all deletions before
# for cut2 allow all deletions after
# for cut1 deletions after, stop if cut2 reached
# for cut2 deletions before, stop if cut1 reached
if cut1_del != (0, 0) and cut2_del != (0, 0):
# todo, this should also handle the straight dropout case w/ no extra deletions
if dropout:
cut1_del_after = 0
cut2_del_before = 0
else:
cut1_del_after = cut1_del[1]
cut2_del_before = cut2_del[0]
cut1 = (cutsite1, cut1_del[0], cut1_del_after)
cut2 = (cutsite2, cut2_del_before, cut2_del[1])
for cutsite, del_before, del_after in [cut1, cut2]:
deleted_bases += [cutsite - i for i in range(del_before)] + [
cutsite + i + 1 for i in range(del_after)
]
for idx, base in enumerate(self.wt_basecalls):
if idx in deleted_bases:
proposal_base = ProposalBase('-', ProposalBase.DELETION,
idx)
else:
proposal_base = ProposalBase(base, ProposalBase.WILD_TYPE,
idx)
for base_index, base_color in enumerate(self.base_order):
proposal_trace.append(self.wt_trace[base_color][idx])
proposal_bases.append(proposal_base)
ep = EditProposal()
ep.sequence_data = proposal_bases
ep.cutsite = cutsite1
ep.cutsite2 = cutsite2
total_deleted = cut1_del[
0] + cut1_del_after + cut2_del_before + cut2_del[1]
total_deleted += -(cutsite2 - cutsite1)
ep.bases_changed = total_deleted
ep.summary = "{}:{}-{}[{}],-{}[{}]".format(
total_deleted, summary_code, cut1_del[0] + cut1_del_after,
label1, cut2_del_before + cut2_del[1], label2)
ep.trace_data = proposal_trace
# insertion case
elif cut1_ins != 0 or cut2_ins != 0:
if dropout:
cut2_ins = 0
cut1 = (cutsite1, cut1_ins)
cut2 = (cutsite2, cut2_ins)
for idx, base in enumerate(self.wt_basecalls):
if idx in deleted_bases:
proposal_bases.append(
ProposalBase('-', ProposalBase.DELETION, idx))
# if base is in range where we need to do insertion
elif idx in [cutsite1, cutsite2]:
for cutsite, insertion_length in [cut1, cut2]:
if cutsite == idx:
# if cutsite != cutsite2:
proposal_bases.append(
ProposalBase(base, ProposalBase.WILD_TYPE,
idx))
for base_index, base_color in enumerate(
self.base_order):
proposal_trace.append(
self.wt_trace[base_color][idx])
# elif dropout:
# proposal_bases.append(ProposalBase('-', ProposalBase.DELETION, idx))
for i in range(insertion_length):
proposal_bases.append(
ProposalBase('n', ProposalBase.INSERTION,
idx))
for base_index, base in enumerate(
self.base_order):
proposal_trace.append(0.25)
else:
proposal_bases.append(
ProposalBase(base, ProposalBase.WILD_TYPE, idx))
for base_index, base_color in enumerate(self.base_order):
proposal_trace.append(self.wt_trace[base_color][idx])
ep = EditProposal()
ep.sequence_data = proposal_bases
ep.cutsite = cutsite1 + cut1_ins
ep.cutsite2 = cutsite1 + cut1_ins + (cutsite2 -
cutsite1) + cut2_ins
ep.bases_changed = cut1_ins + cut2_ins
if dropout:
ep.bases_changed -= (cutsite2 - cutsite1)
ep.summary = "{}:{}+{}[{}],+{}[{}]".format(ep.bases_changed,
summary_code, cut1_ins,
label1, cut2_ins,
label2)
ep.trace_data = proposal_trace
# the intervening sequence is dropped out and no bases inserted or deleted
else:
if dropout:
for idx, base in enumerate(self.wt_basecalls):
if idx in deleted_bases:
proposal_base = ProposalBase('-',
ProposalBase.DELETION,
idx)
else:
proposal_base = ProposalBase(base,
ProposalBase.WILD_TYPE,
idx)
for base_index, base_color in enumerate(
self.base_order):
proposal_trace.append(
self.wt_trace[base_color][idx])
proposal_bases.append(proposal_base)
ep = EditProposal()
ep.sequence_data = proposal_bases
ep.cutsite = cutsite1
ep.cutsite2 = cutsite2
total_deleted = -(cutsite2 - cutsite1)
ep.bases_changed = total_deleted
ep.summary = "{}:{}-0[{}],-0[{}]".format(
total_deleted, summary_code, label1, label2)
ep.trace_data = proposal_trace
else:
# wild type case, use the single edit to model this case
return None
return ep