def test_multiplex_cut():
wt_basecalls = "GCACCTGTCCCCATAGAAATGGCTATGGAAAGCCTTTGGGTTATTTGCG"
"GCACCTGTCC-CCATAGAAATGGCTATGGAAAGCCT-TTGGGTTATTTGCG"
epc = EditProposalCreator(wt_basecalls, use_ctrl_trace=False)
dropout_proposal = epc.multiplex_proposal(10, 35, "test1", "test2", dropout=True)
assert dropout_proposal.sequence == "GCACCTGTCCTTGGGTTATTTGCG"
assert dropout_proposal.summary.startswith('-25:md')
dropout_ins_proposal = epc.multiplex_proposal(10, 35, "test1", "test2", dropout=True, cut1_ins=2, cut2_ins=3)
assert dropout_ins_proposal.sequence == "GCACCTGTCCnnTTGGGTTATTTGCG"
indep_cut = epc.multiplex_proposal(10, 35, "test1", "test2", cut1_del=(4, 0), cut2_del=(0, 3))
assert indep_cut.sequence == "GCACCTCCATAGAAATGGCTATGGAAAGCCTGGTTATTTGCG"
assert indep_cut.summary.startswith('-7:m-4[test1]')
indep_cut2 = epc.multiplex_proposal(10, 35, "test1", "test2", cut1_del=(-4, 0), cut2_del=(3, 0))
assert indep_cut2.sequence == "GCACCTGTCCCCATAGAAATGGCTATGGAAAGTTGGGTTATTTGCG"
def test_human_readable_sequence_multiplex_proposal():
wt_basecalls = ('CCCATAAATCCGCTTTACAGTCACTAGACTAGGACCAATTTCACGACAGGGACTAGACCTTGCTGGATAGCACCTGTCCCCATAGAAATGGCTATGGA'
'AAGCCTTTGGGTTATTTGCGGCACCTGTCCCCATAGAAATGGCTATGGAAAGCCTTTGGGTTATTTGCG')
epc = EditProposalCreator(wt_basecalls, use_ctrl_trace=False)
# test close together dropout default padding
close_together_dropout_proposal = epc.multiplex_proposal(30, 50, 'g1', 'g2', dropout=True)
seq = close_together_dropout_proposal.human_readable_sequence()
assert seq == 'AAATCCGCTTTACAGTCACTAGACT|--------------------|GACTAGACCTTGCTGGATAGCACCTGTCCC'
# test far apart dropout default padding
far_apart_dropout_proposal = epc.multiplex_proposal(30, 90, 'g1', 'g2', dropout=True)
seq = far_apart_dropout_proposal.human_readable_sequence()
assert seq == ('AAATCCGCTTTACAGTCACTAGACT|------------------------------------------------------------|GCTATGGAAAGC'
'CTTTGGGTTATTT')
# test far apart insertion default padding
far_apart_dropout_proposal = epc.multiplex_proposal(30, 90, 'g1', 'g2', dropout=False, cut1_ins=2, cut2_ins=4)
seq = far_apart_dropout_proposal.human_readable_sequence()
assert seq == ('ATCCGCTTTACAGTCACTAGACTnn|AGGACCAATTTCACGACAGGGACTAGACCTTGCTGGATAGCACCTGTCCCCATAGAAATGnnnn|GCTATGGA'
'AAGCCTTTGGGTTATTT')
def _generate_edit_proposals(self):
epc = EditProposalCreator(self.control_sample.primary_base_calls,
use_ctrl_trace=True,
sanger_object=self.control_sample)
proposals = []
# only consider HR for first guide_target
# we have already run homologous_recombination_proposal earlier, so no need to catch exceptions
if self.donor_odn is not None:
cutsite = self.guide_targets[0].cutsite
hr_proposal, changed_bases, odn_start_pos, aln = epc.homologous_recombination_proposal(
cutsite=cutsite, donor_sequence=self.donor_odn)
proposals.append(hr_proposal)
# single cut cases
for guide_target in self.guide_targets:
cutsite = guide_target.cutsite
# deletion case
deletion_befores = list(range(self.indel_max_size + 1))
deletion_afters = list(range(self.indel_max_size + 1))
for deletion_before in deletion_befores:
for deletion_after in deletion_afters:
ep = epc.single_cut_edit_proposal(
cutsite,
guide_target.label,
del_before=deletion_before,
del_after=deletion_after)
proposals.append(ep)
insertions = list(range(self.indel_max_size))
for insertion in insertions:
ep = epc.single_cut_edit_proposal(cutsite,
guide_target.label,
insertion=insertion)
proposals.append(ep)
# multiplex case
# we limit the deletion sizes here
deletion_befores = list(range(5))
deletion_afters = list(range(5))
insertions = list(range(3))
for combo in combinations(self.guide_targets, 2):
guide1 = min(combo, key=lambda x: x.cutsite)
guide2 = max(combo, key=lambda x: x.cutsite)
cutsite1 = guide1.cutsite
cutsite2 = guide2.cutsite
if cutsite1 == cutsite2:
print("Warning: cutsite1 == cutsite2")
continue
label1 = guide1.label
label2 = guide2.label
for cut1_before in deletion_befores:
for cut1_after in deletion_afters:
for cut2_before in deletion_befores:
for cut2_after in deletion_afters:
independent_cut = epc.multiplex_proposal(
cutsite1,
cutsite2,
label1,
label2,
cut1_del=(cut1_before, cut1_after),
cut2_del=(cut2_before, cut2_after))
if independent_cut:
proposals.append(independent_cut)
# dropout case
for cut1_before in deletion_befores:
for cut2_after in deletion_afters:
dropout = epc.multiplex_proposal(cutsite1,
cutsite2,
label1,
label2,
cut1_del=(cut1_before, 0),
cut2_del=(0, cut2_after),
dropout=True)
if dropout:
proposals.append(dropout)
for insertion1 in insertions:
for insertion2 in insertions:
cut_and_insert = epc.multiplex_proposal(
cutsite1,
cutsite2,
label1,
label2,
cut1_ins=insertion1,
cut2_ins=insertion2)
if cut_and_insert:
proposals.append(cut_and_insert)
# dropout insertion case
for insertion in insertions:
dropout_and_insert = epc.multiplex_proposal(cutsite1,
cutsite2,
label1,
label2,
cut1_ins=insertion,
dropout=True)
if dropout_and_insert:
proposals.append(dropout_and_insert)
self.proposals = proposals
def _generate_edit_proposals(self):
epc = EditProposalCreator(self.control_sample.primary_base_calls,
use_ctrl_trace=True,
sanger_object=self.control_sample)
proposals = []
if self.donor_odn is not None:
cutsite = self.guide_targets[0].cutsite
hr_proposal, changed_bases, odn_start_pos, aln = epc.homologous_recombination_proposal(
cutsite=cutsite, donor_sequence=self.donor_odn)
proposals.append(hr_proposal)
# single cut cases
for guide_target in self.guide_targets:
cutsite = guide_target.cutsite
# deletion case
deletion_befores = list(range(self.indel_max_size + 1))
deletion_afters = list(range(self.indel_max_size + 1))
for deletion_before in deletion_befores:
for deletion_after in deletion_afters:
indel_size = -(deletion_before + deletion_after)
if self._should_skip_proposal(indel_size):
continue
ep = epc.single_cut_edit_proposal(
cutsite,
guide_target.label,
del_before=deletion_before,
del_after=deletion_after)
proposals.append(ep)
insertions = list(range(self.indel_max_size + 1))
for insertion in insertions:
if self._should_skip_proposal(insertion):
continue
ep = epc.single_cut_edit_proposal(cutsite,
guide_target.label,
insertion=insertion)
proposals.append(ep)
# multiplex case
# we limit the deletion sizes here
deletion_befores = list(range(5))
deletion_afters = list(range(5))
insertions = list(range(3))
for combo in combinations(self.guide_targets, 2):
guide1 = min(combo, key=lambda x: x.cutsite)
guide2 = max(combo, key=lambda x: x.cutsite)
cutsite1 = guide1.cutsite
cutsite2 = guide2.cutsite
if cutsite1 == cutsite2:
print("Warning: cutsite1 == cutsite2")
continue
label1 = guide1.label
label2 = guide2.label
for cut1_before in deletion_befores:
for cut1_after in deletion_afters:
for cut2_before in deletion_befores:
for cut2_after in deletion_afters:
independent_cut = epc.multiplex_proposal(
cutsite1,
cutsite2,
label1,
label2,
cut1_del=(cut1_before, cut1_after),
cut2_del=(cut2_before, cut2_after))
if independent_cut:
proposals.append(independent_cut)
# dropout case
for cut1_before in deletion_befores:
for cut2_after in deletion_afters:
dropout = epc.multiplex_proposal(cutsite1,
cutsite2,
label1,
label2,
cut1_del=(cut1_before, 0),
cut2_del=(0, cut2_after),
dropout=True)
if dropout:
proposals.append(dropout)
for insertion1 in insertions:
for insertion2 in insertions:
cut_and_insert = epc.multiplex_proposal(
cutsite1,
cutsite2,
label1,
label2,
cut1_ins=insertion1,
cut2_ins=insertion2)
if cut_and_insert:
proposals.append(cut_and_insert)
# dropout insertion case
for insertion in insertions:
dropout_and_insert = epc.multiplex_proposal(cutsite1,
cutsite2,
label1,
label2,
cut1_ins=insertion,
dropout=True)
if dropout_and_insert:
proposals.append(dropout_and_insert)
#removing degenerate proposals
seen = []
self.proposals = list(
filter(
lambda x: seen.append(x.sequence) is None
if x.sequence not in seen else False, proposals))
def test_bad_multiplex():
wt_basecalls = "GCACCTGTCCCCATAGAAATGGCTATGGAAAGCCTTTGGGTTATTTGCG"
epc = EditProposalCreator(wt_basecalls, use_ctrl_trace=False)
with pytest.raises(Exception):
epc.multiplex_proposal(20, 10, "test1", "test2", dropout=True)