def realign_variable(self, germline_gene, match=3, mismatch=-2, gap_open_penalty=22, gap_extend_penalty=1):
'''
Due to restrictions on the available scoring parameters in BLASTn, incorrect truncation
of the v-gene alignment can occur. This function re-aligns the query sequence with
the identified germline variable gene using more appropriate alignment parameters.
Input is the name of the germline variable gene (ex: 'IGHV1-2*02').
'''
# self.germline_seq = self._get_germline_sequence_for_realignment(germline_gene, 'V')
alignment = local_alignment(self.seq.sequence, self.germline_seq,
match=match, mismatch=mismatch,
gap_open_penalty=gap_open_penalty, gap_extend_penalty=gap_extend_penalty)
rc = self.seq.reverse_complement
alignment_rc = local_alignment(rc, self.germline_seq,
match=match, mismatch=mismatch,
gap_open_penalty=gap_open_penalty, gap_extend_penalty=gap_extend_penalty)
if alignment.score > alignment_rc.score:
self._process_realignment(alignment)
else:
self.strand = 'minus'
self.input_sequence = rc
self._process_realignment(alignment_rc)
def _fallback_find_junc_nt_start(self, antibody):
self.fallback_5prime = True
# get the FR3 nt sequence of the IMGT gapped germline
germ_fr3_sequence = antibody.v.imgt_germline.gapped_nt_sequence[196:309].replace('.', '')
antibody.log('GERM FR3 SEQUENCE:', germ_fr3_sequence)
# find the start of the junction (immediately after the end of FR3)
aln = local_alignment(antibody.oriented_input, germ_fr3_sequence)
antibody.log(' QUERY: ', aln.aligned_query)
antibody.log(' ', aln.alignment_midline)
antibody.log('GERM FR3:', aln.aligned_target)
fr3_end = aln.query_end + (len(germ_fr3_sequence) - aln.target_end)
junc_start_codon = antibody.oriented_input[fr3_end:fr3_end + 3]
antibody.log('JUNC START:', junc_start_codon, codons[junc_start_codon], fr3_end)
return fr3_end
def _fallback_find_junc_nt_end(self, antibody):
self.fallback_3prime = True
# need to find the start of FR4 in the IMGT germline sequence
end_res = 'W' if antibody.chain == 'heavy' else 'F'
for i, res in enumerate(antibody.j.imgt_germline.ungapped_aa_sequence):
if res == end_res and end_res not in antibody.j.imgt_germline.ungapped_aa_sequence[i + 1:]:
fr4_nt_start_pos = (antibody.j.imgt_germline.coding_start - 1) + (i * 3)
break
germ_fr4_sequence = antibody.j.imgt_germline.gapped_nt_sequence[fr4_nt_start_pos:]
# find the end of the junction (end of the first codon of FR4)
aln = local_alignment(antibody.oriented_input, germ_fr4_sequence)
fr4_start = aln.query_begin - aln.target_begin
junc_end_codon = antibody.oriented_input[fr4_start:fr4_start + 3]
antibody.log('JUNC END:', junc_end_codon, codons[junc_end_codon], fr4_start)
return fr4_start + 3
def realign_germline(self, antibody, query_start=None, query_end=None):
'''
Due to restrictions on the available scoring parameters in BLASTn, incorrect truncation
of the v-gene alignment can occur. This function re-aligns the query sequence with
the identified germline variable gene using more appropriate alignment parameters.
Args:
oriented_input (str): the raw input sequence, correctly oriented
query_start (int): 5' position in `oriented_input` at which the sequence
should be truncated prior to alignment with the germline sequence.
query_end (int): 3' position in `oriented_input` at which the seqeunce
should be truncated prior to alignment with the germline sequence
'''
oriented_input = antibody.oriented_input
germline_seq = self._get_germline_sequence_for_realignment()
aln_params = self._realignment_scoring_params(self.gene_type)
# if the alignment start/end positions have been annotated by the assigner,
# force re-alignment using those parameters
if all([x is not None for x in [self.query_start,
self.query_end,
self.germline_start,
self.germline_end]]):
query = oriented_input.sequence[self.query_start:self.query_end]
germline = germline_seq[self.germline_start:self.germline_end]
alignment = global_alignment(query, germline, **aln_params)
# use local alignment to determine alignment start/end positions if
# they haven't already been determined by the assigner
else:
query = oriented_input.sequence[query_start:query_end]
alignment = local_alignment(query, germline_seq, **aln_params)
if alignment:
self._process_realignment(antibody, alignment, query_start)
else:
antibody.log('GERMLINE REALIGNMENT ERROR')
antibody.log('REALIGNMENT QUERY SEQUENCE:', query)
antibody.log('QUERY START:', query_start)
antibody.log('QUERN END:', query_end)
def assign_d(seq, species):
'''
Identifies the germline diversity gene for a given sequence.
Alignment is performed using the ssw_wrap.Aligner.align function.
Input is a junction sequence (as a string) and the species of origin.
Output is a DiversityResult object.
'''
mod_dir = os.path.dirname(os.path.dirname(os.path.abspath(__file__)))
db_file = os.path.join(mod_dir, 'ssw/dbs/{}_D.fasta'.format(species.lower()))
db_handle = open(db_file, 'r')
germs = [Sequence(s) for s in SeqIO.parse(db_handle, 'fasta')]
rc_germs = [Sequence(s.reverse_complement, id=s.id) for s in germs]
germs.extend(rc_germs)
db_handle.close()
alignments = local_alignment(seq, targets=germs,
gap_open_penalty=20, gap_extend_penalty=2)
alignments.sort(key=lambda x: x.score, reverse=True)
try:
return blast.DiversityResult(seq, alignments[:5])
except IndexError:
return None
def gapped_imgt_realignment(self):
'''
Aligns to gapped IMGT germline sequence. Used to determine
IMGT-formatted position numberings so that identifying
antibody regions is simplified.
'''
self.imgt_germline = get_imgt_germlines(species=self.species,
gene_type=self.gene_type,
gene=self.full)
query = self.germline_alignment.replace('-', '')
aln_params = self._realignment_scoring_params(self.gene_type)
aln_params['gap_open'] = -11
aln_matrix = self._get_gapped_imgt_substitution_matrix()
self.imgt_gapped_alignment = local_alignment(query,
self.imgt_germline.gapped_nt_sequence,
matrix=aln_matrix,
**aln_params)
self.alignment_reading_frame = ((2 * (self.imgt_gapped_alignment.target_begin % 3)) % 3) + (self.imgt_germline.coding_start - 1) # IMGT coding start is 1-based
self.coding_region = self._get_coding_region()
self.aa_sequence = self._get_aa_sequence()
try:
self._imgt_numbering()
except:
self.exception('IMGT NUMBERING', traceback.format_exc(), sep='\n')
def _find_junction_nt_end(self, vdj):
fr4 = vdj.j.regions.nt_seqs['FR4'][3:]
aln = local_alignment(fr4, vdj.vdj_nt)
if aln:
return aln.target_begin
def _find_junction_nt_start(self, vdj):
fr3 = vdj.v.regions.nt_seqs['FR3'][:-3]
aln = local_alignment(fr3, vdj.vdj_nt)
if aln:
return aln.target_end + 1
def _get_d_start_position_nt(self, vdj):
a = local_alignment(self.d_nt, self.cdr3_nt,
gap_open_penalty=22, gap_extend_penalty=1)
d_start = a.target_begin
return d_start
def _get_isotype_query_region(self, antibody):
aln = local_alignment(antibody.vdj_nt, antibody.oriented_input)
return antibody.oriented_input[aln.target_end:]
def _get_alignments(self, antibody, isotype_seqs):
query_region = self._get_isotype_query_region(antibody)
alignments = local_alignment(query_region, targets=isotype_seqs,
gap_open_penalty=22, gap_extend_penalty=1)
return sorted(alignments, key=lambda x: x.score, reverse=True)
def _get_isotype_query_region(self, vdj):
aln = local_alignment(vdj.vdj_nt, vdj.raw_input)
return vdj.raw_input[aln.target_end:]
def _get_alignments(self, vdj, isotype_seqs):
query_region = self._get_isotype_query_region(vdj)
alignments = local_alignment(query_region, targets=isotype_seqs,
gap_open_penalty=22, gap_extend_penalty=1)
return sorted(alignments, key=lambda x: x.score, reverse=True)
