def add_primers_to_alignment(primers, alignment, reverse=False):
alignment = alignment.clone()
ali_len = alignment.get_alignment_length()
if reverse:
for p in primers:
alignment.append(
copy_attrs(p.master_sequence,
p.master_sequence.reverse_complement()) + '-' *
(ali_len - len(p)))
else:
for p in primers:
alignment.append(p.master_sequence + '-' * (ali_len - len(p)))
return AlignmentUtils.align(alignment)
def find_primers(segments,
foi,
pfinder_args,
reverse=False,
qualifier='ugene_name'):
genes = []
for s in segments:
i = get_feature_indexes_by_qualifier(s, qualifier, foi)
if not i: continue
genes.append(s.features[i[0]].extract(s))
ali = AlignmentUtils.align(genes)
if not ali: return None
primer_alis = PrimerFinder.find_specific_primers(ali,
reverse=reverse,
**pfinder_args)
return PrimerFinder.compile_primers(primer_alis, foi + '_', reverse), ali
def add_pairs_to_alignment(pairs, alignment):
added = set()
alignment = alignment.clone()
ali_len = alignment.get_alignment_length()
for i, p in enumerate(pairs):
if p.forward.id not in added:
alignment.append(p.forward.master_sequence + '-' *
(ali_len - len(p.forward)))
added.add(p.forward.id)
if p.reverse.id not in added:
alignment.append(
copy_attrs(p.reverse.master_sequence,
p.reverse.master_sequence.reverse_complement())
+ '-' * (ali_len - len(p.reverse)))
added.add(p.reverse.id)
return AlignmentUtils.align(alignment)
def _main(self):
min_prod = 400
silva_db = '/home/allis/Documents/INMI/SILVA-DB/SILVA_123_SSURef_Nr99_tax_silva.fasta'
alifile = '/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.aln.fasta'
add_filename = FilenameParser.strip_ext(alifile)+'.with_additions.fasta'
outgroups = ['Thermococcus_chitonophagus', 'SMTZ1-55', 'contig72135_1581_sunspring_meta']
add = ['KF836721.1.1270','EU635905.1.1323']
exclude = []#['Thermococcus_chitonophagus', 'SMTZ1-55', 'BA1-16S', 'contig72135_1581_sunspring_meta']
#load alignment
if os.path.isfile(add_filename):
alifile = add_filename
add_filename = ''
with user_message('Loadding initial alignment...', '\n'):
orig_ali = AlignmentUtils.load_first(alifile)
if not orig_ali: return 1
#load homologs
if add_filename:
with user_message('Loadding additional sequences...', '\n'):
add_seqs = []
db = SeqView()
if db.load(silva_db):
for sid in add:
seq = db.get(sid)
if seq: add_seqs.append(seq)
else: print '%s not found in %s' % (sid, silva_db)
#realign data if needed
if add_seqs:
with user_message('Realigning data...', '\n'):
add_filename = FilenameParser.strip_ext(alifile)+'.with_additions.fasta'
AlignmentUtils.align(list(orig_ali)+add_seqs, add_filename)
orig_ali = AlignmentUtils.load_first(add_filename)
if not orig_ali: return 2
#process the alignment
ali = orig_ali.remove(*exclude).trim()
for out in outgroups:
if not ali.index(out):
print '%s not found in the alignment' % out
return 3
ali.sort(key=lambda r: 'zzzzzzzz' if r.id in outgroups else r.id)
ali_len = ali.get_alignment_length()
AlignmentUtils.save(ali, '/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.aln.trimmed.fasta')
args = dict(plen = (20,40),
max_mismatches = 8,
min_match_mismatches = 1,
first_match_mismatches = 1,
first_may_match = 1,
AT_first=True,
outgroup=len(outgroups))
fprimers = self._find_primers(ali, **args)
rprimers = self._find_primers(ali.reverse_complement(), **args)
pairs = []
for i, (fs, fp) in enumerate(fprimers):
start = fs
fprimer = Primer.from_sequences(fp[:-1], 1, 'SSBaF%d' % fs)
for _j, (rs, rp) in enumerate(rprimers):
end = ali_len-rs
if end-start <= min_prod: continue
pairs.append((fprimer, Primer.from_sequences(rp[:-1], 1, 'SSBaR%d' % (ali_len-rs+1))))
if not pairs:
print '\nNo suitable primer pairs found'
return 3
added = set()
for i, (fp, rp) in enumerate(pairs):
print '\npair %d' % (i+1)
print '%s: %s' % (fp.id, fp)
print '%s: %s' % (rp.id, rp)
if fp.id not in added:
orig_ali.append(fp.master_sequence+'-'*(orig_ali.get_alignment_length()-len(fp)))
added.add(fp.id)
if rp.id not in added:
orig_ali.append(copy_attrs(rp.master_sequence,
rp.master_sequence.reverse_complement())+
'-'*(orig_ali.get_alignment_length()-len(rp)))
added.add(rp.id)
print
orig_ali = AlignmentUtils.align(orig_ali)
AlignmentUtils.save(orig_ali, '/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.with_primers.aln.fasta')
print 'Done'
def _main(self):
query = simple_rec(
'AAACTGGGGCTAATACCCGATGGGTGAGGAGGCCTGGAATGGTTCTTCACCGAAAAGACGTTGAGACCATGCTTTTCAACGTTGCCTAAGGATGGGGCCGCGTCCGATCAGGTTGTTGGTGGGGTAACGGCTCACCAAGCCTATAACCGGTACGGGCCGTGGGAGCGGAAGCCCGGAGATGGGCACTGAGACAAGGGCCCAGGCCCTACGGGGCGCAGCAGTCGCGAAAACTCCGCAATGCGCGAAAGCGTGACGGGGCTACCCCGAGTGCCGTCCGCTGAGGATGGCTTTTCCCCGGTGTAATGAGCCTGGGGAATAAGGAGAGGGCAAGCCTGGTGTCAGCCGCCGCGGTAATACCAGCTCTCCGAGTGGTAGGGATGATTATTGGGCTTAAAGCGTCCGTAGCCAGCCCGGCAAGTCTCCCGTTAAATCCAGCGACCTAATCGTTGGGCTGCGGAAGATACTGTTGGGCTAGGGGGCGGGAGAGGCCGACGGTATTCCCGGGGTAGGGGTGAAATCCTATAATCCTGGGAGGACCACCAGTGGCGAAGGCTGTCGGCTAGAACGCGCTCGACGGTGAGGGACGAAAGCTGGGGGAGCGAACTGGATTAGATACCCGGGTAGTCCCAGCTGTAAACGATGCGGGCTAGGTGTTGGGGTGGCTACGAGCCACCTCAGTGCCGCAGGGAAGCCATTAAGCCCGCCGCCTGGGAAGTACGGCCGCAAGGCTGAAACTTAAAGGAATTGGCGGGGGAGCACCACAAGGCGTGAAGCTTGCGGTTTAATTGGAGTCAACGCCGGGAACCTTACCGGGGGCGACAGCAGGATGAGGGCCAGATTGAAGGTCTTGCTTGACAAGCTGAGAGGAGGTGCATGGCCGTCGCCAGTTCGTGCCGTGAGGTGTCCTGTTAAGTCAGGCAACGATCGAGACCCGCACCCTTAGTTGCAACCCCTGCGGAACCCGCAGGGGGCACACTACGGGAACTGCCGCCGATAAGGCGGAGGAAGGAGCGGGCCACGGCAGGTCAGTATGCCCCGAATCCCCCGGGCCACACGCGAGCTGCAATGGCAGAGACAATGGGTTCCAACCTTGAAAGAGGGAGGTAATCCCTAAACCCTGCCTCAGTTGGGATCGAGGGCTGCAACCCGCCCTCGTGAACATGGAATGCCTAGTAATCGCGTGTCATCATCGCGCGGTGAATACGTCCCCGCTCCTTGCACACACCGCCCGTCGCTCCATCCGAGTGGGGTTTGGGTGAGGCGTGGTCTGTTGGCCGCGTCGAATCTAGGCTTCGCGAGGAGGGAGAAGTCGTAACAAGGTGGCCGTAGGGGAACCTGCGGCCGGATCACCTCCT',
'BA2-16S')
suns_db = '/home/allis/Documents/INMI/SunS-metagenome/BlastDB-big/sunspring_meta'
silva_db = '/home/allis/Documents/INMI/SILVA-DB/SILVA_123_SSURef_Nr99_tax_silva'
additions = [
simple_rec(
'AAACTGGGGCTAATCCCCCATAGGCCTGGGGTACTGGAAGGTCCCCAGGCCGAAAGGG------GACCGTA-----AGGTCCCGCCCGAGGATGGGCCGGCGGCCGATTAGGTAGTTGGTGGGGTAACGGCCCACCAAG--CCGAAGATCGGTACGGGCC-GTGAGAGCGGGAGCCCGGAGATGGACA---CTGAGACACGGGTCCAGGCCCTACGGGGCGCAGCAGGCGCGAAACC-TCCGCAATGCGGGAAACCGCGACGGGGGGACCCCCAGTGCCGTGCCTCTGGC-----ACGGCTTTTCCGGAGTG-TAAAAAGCTCCGGGAATAAGGGCTGGGCAAGGCCGGTGGC-AGCCGCCGCGGTAATACCGGCGGCCCGAGTGGTGGCCACTATTATTGGGCCTAAAGCGGCCGTAGCCGGGCCCGTAAGTCCCTGGCG-AAATCCCACGGCTCAACCGTGGGGCTCGCTGGGGATACTGCGG-GCCTTGGGACCGGGAGAGGCCGGGGGTACC-CCCGGGGTAGGGGTGAAATCCTATAATCCCGGGGGGACCGCCAGT-GGCGAAGGCGCCC--GGCTGGAACGGGTCCGACGGTGAGGGCCGAAGGCC-AGGGGAGCGAACCGGATTAGATACCCGGGTAGTCCTGGCTGTAAAGGATGCGGGCTAGGTGTCGGGCGAG-CTTCGAGCTCGC-CCGGTGCCGTAGGGAAGCCGTTAAGCCCGCCGCC-TGGGGAGTACGGCCGCAAGGCT-GAAACTTAAAGGAATT-GGCGGGGGAGC-ACTACAAGGGGTGGAGCGTGCGGTTTAATTGGATTCAACGCCGGGAACCTCACCGGGGGCGACGGCAGGATGAA-GGCCAGGCTGAAGGTCTTGCCGGACGCGCCGAGAGGAG-----------------------------------GTGCATGGCCGCCGTCAGCTCGTACCGTGAGGCGTCCA-CTTAAGTGTGGTAACGAGCGAGACCCGC--GCCCCCAGTTGCCAGTCCCTCCCGCTGGGA---GGGAGGC-ACTCTGGGGGG-ACTGCCGGCGAT-AAGCCGGAGGAAGGGGCGGGCGACGGTAGGTCAGTATG-CCCCGAAACCC-CCGGGCT-ACACGCGCGCTACAATGGGCGGGACAATGGGA-CCCGACCCCGAAAGGGGAAGGGAATCCCCTAAACCCGCCCTCAGTTCGGATCGCGGGCTG-CAACTCGCCCGCGTGAAGC-TGGAAT-CCCTAGTACCCGCGCGTCATCATCGCGCGGCGAATACGTCCCTGCTCCTTGCACACACCGCCCGTCACTCCACCCGAG-CGGGGCCC-GGGTGAGGCCCGATCTCCTTCGGGAGGTCGGGTCGAGCCTGGGCTC-CGTGAGGGGGG-AGAAGTCGTAACAAGGTAGCC------------------------------'
.replace('-', ''), 'Thermococcus_chitonophagus'),
simple_rec(
'AAACTGGGATTAATACCCACTAAATGATAATACCTGGAATGGCTTATCATTGAAAGAC-TCTGGAAACATGCTTC-CAGCGTCGCCCAAGG-------------------------------------------------------------------------------GGAGCCCGGAGATGGAAA---CTGAGACAAGGTTCCAGGCCCTACGGGGCGCAGCAGGCGCGAAACC-TCCACAATGCGCGAAAGCGTGATGGGGTTATCCCGAGTGCCGTCCGATGAGG-----ATGGCTTTTCCTCGGTG-TAAGGATCCGAGGGAATAAAGGGGGGGCAAGACTGGTGTC-AGCCGCCGCGGTAATACCAGCTCCCTGAGTGGTAAGGACGATTATTTGGCCTAAAGCGTCCGTAGCCGGCTTATCAAGTCTCTTGTT-AAACCCAGTGATTCAATCATTGACCT-GCAAGAGATACTGTTA-TGCTAGAGGACGGGAGAGGTCGACGG---------GGGTAGGGGTGAAATCCTATAATCCTTGGAGGACCACCAGT-GGCGAAGGCGGTC--GACTAGAACGTGCCTGACGGTGAGGGACGAAAGCT-GGGGGAGCGAACCGGATTAGATACCCGGGTAGTCCCAGCTGTAAACGATGCGGGCTAGGTGTTGGGGTAG-CTACGAGCTACT-CCAGTGCCGCAGAGAAGTTGTTAAGCCCGCCGCC-TGGGGAGTACGGCCGCAAGGCT-GAAACTTAAAGGAATT-GGCGGGGGAGC-ACCACAAGGGGTGAAGGCTGCGGTTTAATTGGAGTCAACGCCGGGAACCTTACCGGGGCTGACAGCAGAGTGAA-GGCCAGACTGAAGATCTTGCCAGACAAGCTGAGAGGAGGTGCATGAAGATCTTGCCAGACAAGCTGAGAGGAGGTGCATGGCCGTCGCCAGTTCGTGCCGTGAGGTGTCCT-GTTAAGTCAGGCAACGAACGAGACCCCC--ACTGTTAGTTGCCAGCGAATTCCAACGGAAT--GTCGGGC-ACACTAACAGG-ACTGCCACCGAT-AAGGTGGAGGAAGGAGGGGGCAACGGCAGGTCAGTATG-CCCC--------------------------------------------------------------------------------------------------------------GAACTCGCCCTCATGAACA-TGGAAT-CCCTAGTAACCGCGTGTCATCATCGCGCGGTGAATACGTCCCCGCTCCTTGCACACACCGCCCGTCGCTCCATCCAAG-TCGGGTCT-AGATGAGGCGCAGTCTTCT-----TGGCTACGTCGAATCTGGGTTC-GGTGAGGGGGG-AGAAGTCGTAACAAGGTGGCCGTAGGGGAACCTGCGGCCGGATCACCTCCT'
.replace('-', ''), 'SMTZ1-55'),
simple_rec(
'ACTCCGGTTGATCCTGCCGGACCCCACTGCTATCGGGGTAGGACTTAACCATGCGAGTTGTGCGTCCCCAAGCCATGGTGGGGGCGCGGCATACGGCTCAGTAACACGTGGCTAACCTAGCCTTTGGACGGGGACAACCCCGGGAAACTGGGGCTAATCCCCGATGGGTGGGAAGGCCTGGAATGGTTTCCCACCGAAAGGGCGTCTGAACCATGCTTCAGGCGTTGCCGAAGGATGGGGCCGCGGCCGATCAGGTTGTTGGTGAGGTAACGGCTCACCAAGCCTATAACCGGTACGGGCCGTGAGAGCGGGAGCCCGGAGATGGGCACTGAGACAAGGGCCCAGGCCCTACGGGGCGCAGCAGGTGCGAAAACTCCGCGATGCGCGAAAGCGTGACGGGGCTATCCCGAGTGCCGTCCGCTGAGGATGGCTTTTCCCCGGTGTAGGGAGCCGGGGGAATAAGGAGAGGGCAAGTCTGGTGTCAGCCGCCGCGGTAATACCAGCTCTCCGAGTGGTGGGGACAATTATTGGGCTTAAAGCGTCCGTAGCCGGCCCATCAAGTCTCTTGTTAAATCCAGCGATCCAATCGCTGGACTGCGGGAGATACTGCTGGGCTAGGGGGCGGGAGAAGCCGATGGTATTCTCGGGGTAGGGGTGAAATCCTATAATCCCGGGAGGACCACCAGTGGCGTAGGCGGTCGGCTAGAACGCGCCCGACGGTGAGGGACGAAAGCTGGGGGAGCGAACCGGATTAGATACCCGGGTAGTCCCAGCCGTAAACGATGCGGGCTAGGTGTTGGGGTGGCTACGAGCCACCCCAGTGCCGCATGGAAGCAATTAAGCCCGCCGCCTGGGGAGTACGGCCGCAAGGCTGAAACTTAAAGGAATTGGCGGGGGAGCACCACAAGGGGTGAAGCTTGCGGTTTAATTGGAGTCAACGCCGGGAAAGGAACAGCGTTTTGTTGTTCCTCTGGATACCTTACCGGGGGCGACAGCAGGATGAAGGCCAGATTGAAGGTCTTGCTGGACGAGCTGAGAGGAGGTGCATGGCCGTCGCCAGTTCGTGCCGTGAGGTGTCCTGTTAAGTCAGGTAACGATCGAGACCCACACCCCCAGTTGCTACCTCTTCGGAGGGCACTCTAGGGGTACTGCCGCCGATAAGGCGGAGGAAGGAGTGGGCCACGGCAGGTCAGTATGCCCCGAATCCCCCGGGCCACACGCGAGCTGCAATGGCAAGGACAATGGGTTCTGACCCCGAGAGGGGAAGGTAATCCCGAAACCCTGCCTCAGTTGGGATCGAGGGCTGAAACCCGCCCTCGTGAACATGGAATCCCTAGTAATCGCGGGTCACCAGCCCGCGGTGAATACGTCCCTGCTCCTTGCACACACCGCCCGTCGCTCCATCCGAGTGGGGTTTAGGTGAGGCGTGGTCCTTGTGGCTGTGTCGAATCTAGGCTTCGCGAGGAGGGAGAAGTCGTAACAAGGTGGCCGTAGGGGAACCTGCGGCCGGATCACCTC',
'BA1-16S'),
simple_rec(
'CTGGTGGAAATATAGAAGAGGCCAAATCCGGGGTTCAGGCCGCCCGGGGTAATTACCCGTTGTCGGAGTGGGGGGGGGACGCTATTGGGGCTTAAGCCATCGTTAGCCCGTTTGACCAGGTCTCTTGTTAAATCAGGCGGATTTATTGGTCGATTGCAGGAGATTATGTTCGTCTTAGGGGCCGGAGGAGTCAACAGTATTCCCGGGGTAGGAGTGAATGCCTATATTCCCGGAGGTACCACCAGTGGGGACGCCGTTGGTATAGAACGCGCCGGCCGGTGATGGAATGAAAGTGAGGGAACCGACCCGAATTAGATACCGGGGTATTGCTACCGTTAACCGATGCAGCTTAGGTGTTCGGGTGGTTACTAGCCATTCGAGTGCGCCAGGGAAGCTGTCAGGCTTACCGCTTGGGAAGTGCGGCTGCAGGGCCAAAACTTAAGGAAATCGCCGGGGAAGCACCCCAGGGGGTGAAGCTTGCGCTTTAATGGAATTCACCGCGGTAATTCTCACCGGGGGAGCCACCAGGAGGAAAGCCAGATTAAAGTTCTTGTTGGCGGAGTGGAGAGGAGGTGCATGCCGTTCGCCAGTTCTTCCCGGGAGGTTCTTGTTAGTTCAGCCACCGATGAGGACCGCCATCCCCTGTTGTTATTGGCCTTGCGCCAGGCACACTGGGGAGACCGCCGCCGATAAGGCGGAGGAAGGAGCGGGCCACGGCAGGTCAGTATGCCCCGAATCCCCCGTCCACACGCGAGGGGCAATG',
'155a'),
simple_rec(
'CAAGTCCTATAACCGGTACGGGCCGTGGGAGCGGTAGCCCGGAGATGGGCACTGAGACAAGGGCCCAGGCCCTACGGGGCGCAGCAGTCGCGAAACCTCCGCAATACGCGAAAGCGTGACGGGGTCATCCCGAGTGCCGTCCGCTGAGGATGGCTTTTCCCCAGTGTAGACAGCTGGGGGAATAAGGAGAGGGCAAGTCGGGTGTCAGCCGCCGCGGTAATACCCGCTCTCCGAGTGGTGGGGACGCTTATTGGGCCTAAAGCATCCGTAGCCGGCTGGACAAGTCCCCTGTTAAATCCAGCGATTTAATCGTTGGACTGCGGGGGATACTGTCCGGCTAGGGGGCGGGAGAGGCCGACGGTATTTCCGGGGTAGGGGTGAAATCCTATAATCCCGGGAGGACCACCAGTGGCGAAGGCTGTCGGCTAGAACGCGCCCGACGGTGAGGGATGAAAGCTGGGGGAGCGAACCGGATTAGATACCCGGGTAGTCCCAGCCGTAAACGATGCAGGCTAGGTGTTCGGGTGGCTACGTGCCACTCGAGTGCCGCAGGGAAGCTGTTAAGCCTGCCGCCTGGGGAGTACGGCCGCAAGGCTGAAACTTAAAGGAATTGGCGGGGGAGCACCACAAGGGGTGAAGCTTGCGGTTTAATTGGAGTCAACGCCGGAAATCTCACCGGGGGAGACAGCAGGATGAAAGCCAGATTAAAGGTCTTGCTAGACGAGCTGAGAGGAGGTGCATGGCCGTCGCCAGTTCGTGCCGTGAGGTGTCCTGTTAAGTCAAGGCAACGATCGAGACTCGCATCCTCTGTTGCTACTACCCTTGCGCCAAGGCACACTGGGGGAGACCGCCGCTCGATAAGGCGGAAGGAAGGAGCGGCCCACGGCAGTCAGTATGCCCCGAATTCCCTCGGCCACACGCAAGCTGCAATG',
'156a'),
simple_rec(
'GGGGATCGGGGCATACTGACCTGCCGTGGCCCGCTCCTTCCTCCGCCTTATCGGCGGCGGTCTCCCCAGTGTGCCTGGCGCAAGGGCAGTAACAACAGGGGATGGGGGTCTCGATCGGTGGCTGGCTTAACAGGAAACCTCACGGGACGAACTGGCGAACGGCATGGACCTTCTCTCAACTTGGCTAAGAAGAACTTTAATCTGGCTTTCATTCTGGTGGCTTCCCCGGTGAGAATTCCGGCGGTGACTCCCAATAAAACGCAAGCTTCACCCCTTGGGGTGGTTCCCCGGCCATTTCTTTAAGGTTCAAGCTTTGCGGCGGTATTCCCAAGCGGCAAGGTTAACAGCTTCCCTGCCGCACTCGAGTGGCACGTAACCACCCGAACAACTAACCTGCATCCGTTACCGGTTGGACTAACCCGGTATCTAATCCGGGTCGCTCCCCCAGCCTTCATTCCTTCACCGTCCGGCGCGGTTCTAAGCGACCGGCTTTCGCACTTGTGGTTCCTCCCGGGGATTATAAGAATTCACCCCTACCCCGGAAATTACGGTCCGGCTCCTCCGGCCCCTAACCCGACACGTAATCCCCCGCCAGTTCAACCGATTAAATCCGCTTGAATTTAACAAGGGGGACCTTGTCCAGCCGGCCTACGGATGCTTTAAGGCCCAATAAGCCGTCCCCACCACTCCGAGAGCGGGTAATAACCGCGGCCGGCCTGACAACCGACCTGGCCTCTCCTAAATCCCCCAGCTGTTCACACTTGGGAAAGGGCATTCCTCAGCGAACGGCACTTCGGGATGAACCCGTCACGCTTTCGCGTAATTGCGGGAAGGTTTCGCGAACTGCTGCGCCCCGTAAAGGCCTGGGTCCTTGTGTCTCAAATTGCCCCATCTCCGGGCTATACGCTCTCCACGGGCCCGTACC',
'157a')
]
#prepare filter
filt = BlastFilter(lambda a, r: a.hsps[0].align_length > 1100)
filt.AND = BlastFilter(
lambda a, r: all(hsp.score > 500 for hsp in a.hsps))
filt.AND.AND = BlastFilter(lambda a, r: all(
hsp.identities / float(hsp.align_length) > 0.8 for hsp in a.hsps))
#make ring-blast
blast = BlastCLI(self.abort_event)
orig_seqs = blast.ring_blast(query, suns_db, 100, filt, 3)
if not orig_seqs:
print 'No blast results.'
return 1
nseqs = len(orig_seqs)
print 'RingBlast to:\n%s\nreturned %d sequences.\n' % (suns_db, nseqs)
#save an initial alignment
self.fix_ids(orig_seqs)
alifile = '/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.aln.fasta'
with user_message('Aligning retrieved sequences...', '\n'):
if not AlignmentUtils.align(orig_seqs + [query] + additions,
outfile=alifile):
return 3
#search for additional homologs
add_seqs = blast.ring_blast(orig_seqs, silva_db, 100, filt, 0)
if add_seqs:
self.fix_ids(add_seqs)
print 'RingBlast to:\n%s\nreturned %d additional sequences.\n' % (
silva_db, len(add_seqs))
#build an alignment
seqs = orig_seqs + add_seqs + [query] + additions
alifile = '/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.big.aln.fasta'
with user_message('Aligning retrieved sequences...', '\n'):
if not AlignmentUtils.align(seqs, outfile=alifile): return 3
#build a tree
treefile = '/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.big.aln.tre'
if not PhyloUtils.build_fast_tree(alifile, treefile): return 4
#annotate the tree
if False:
with open(
'/home/allis/Documents/INMI/16S/SSBaF4-SSBaR4-1_243072232-iPCR-report.txt'
) as inp:
# SSBaF4-SSBaR4_65397396-iPCR-report.txt
sids = set()
len_re = re.compile(r'(\s|^)(\d+)(\sbp|\\s*:)?', re.MULTILINE)
entry = False
cur_sid = None
cur_len = -1
for l in inp:
if l == '========= histograms and electrophorograms of PCR products of each hit =========':
break
if l.startswith('---'):
entry = False
if cur_sid and cur_len > 0 and abs(cur_len - 920) < 60:
sids.add(cur_sid)
cur_sid = None
cur_len = -1
continue
if entry or '#' in l:
entry = True
plen = len_re.search(l)
if plen: cur_len = int(plen.group(2))
sid = BlastID.extract(l)[0]
if sid: cur_sid = sid
organisms = Organisms.from_records(seqs)
if PhyloUtils.annotate_tree(
treefile,
organisms,
reroot_at='Thermococcus_chitonophagus',
# beautify_leafs=True,
# collapse_taxa=['miscellaneous crenarchaeotic group', 'thaumarchaeota'],
# collapse_last=True,
# collapse_hard=True,
# mark_leafs=sids,
mark_leafs=[r.id for r in orig_seqs + [query] + additions],
lineage_colors={
'miscellaneous crenarchaeotic group': (0, 0, 255),
'thaumarchaeta': (255, 0, 0)
},
top_lineage=Lineage('archaea')):
return 0
return 2
def _main(self):
query = simple_rec('AAACTGGGGCTAATACCCGATGGGTGAGGAGGCCTGGAATGGTTCTTCACCGAAAAGACGTTGAGACCATGCTTTTCAACGTTGCCTAAGGATGGGGCCGCGTCCGATCAGGTTGTTGGTGGGGTAACGGCTCACCAAGCCTATAACCGGTACGGGCCGTGGGAGCGGAAGCCCGGAGATGGGCACTGAGACAAGGGCCCAGGCCCTACGGGGCGCAGCAGTCGCGAAAACTCCGCAATGCGCGAAAGCGTGACGGGGCTACCCCGAGTGCCGTCCGCTGAGGATGGCTTTTCCCCGGTGTAATGAGCCTGGGGAATAAGGAGAGGGCAAGCCTGGTGTCAGCCGCCGCGGTAATACCAGCTCTCCGAGTGGTAGGGATGATTATTGGGCTTAAAGCGTCCGTAGCCAGCCCGGCAAGTCTCCCGTTAAATCCAGCGACCTAATCGTTGGGCTGCGGAAGATACTGTTGGGCTAGGGGGCGGGAGAGGCCGACGGTATTCCCGGGGTAGGGGTGAAATCCTATAATCCTGGGAGGACCACCAGTGGCGAAGGCTGTCGGCTAGAACGCGCTCGACGGTGAGGGACGAAAGCTGGGGGAGCGAACTGGATTAGATACCCGGGTAGTCCCAGCTGTAAACGATGCGGGCTAGGTGTTGGGGTGGCTACGAGCCACCTCAGTGCCGCAGGGAAGCCATTAAGCCCGCCGCCTGGGAAGTACGGCCGCAAGGCTGAAACTTAAAGGAATTGGCGGGGGAGCACCACAAGGCGTGAAGCTTGCGGTTTAATTGGAGTCAACGCCGGGAACCTTACCGGGGGCGACAGCAGGATGAGGGCCAGATTGAAGGTCTTGCTTGACAAGCTGAGAGGAGGTGCATGGCCGTCGCCAGTTCGTGCCGTGAGGTGTCCTGTTAAGTCAGGCAACGATCGAGACCCGCACCCTTAGTTGCAACCCCTGCGGAACCCGCAGGGGGCACACTACGGGAACTGCCGCCGATAAGGCGGAGGAAGGAGCGGGCCACGGCAGGTCAGTATGCCCCGAATCCCCCGGGCCACACGCGAGCTGCAATGGCAGAGACAATGGGTTCCAACCTTGAAAGAGGGAGGTAATCCCTAAACCCTGCCTCAGTTGGGATCGAGGGCTGCAACCCGCCCTCGTGAACATGGAATGCCTAGTAATCGCGTGTCATCATCGCGCGGTGAATACGTCCCCGCTCCTTGCACACACCGCCCGTCGCTCCATCCGAGTGGGGTTTGGGTGAGGCGTGGTCTGTTGGCCGCGTCGAATCTAGGCTTCGCGAGGAGGGAGAAGTCGTAACAAGGTGGCCGTAGGGGAACCTGCGGCCGGATCACCTCCT',
'BA2-16S')
suns_db = '/home/allis/Documents/INMI/SunS-metagenome/BlastDB-big/sunspring_meta'
silva_db = '/home/allis/Documents/INMI/SILVA-DB/SILVA_123_SSURef_Nr99_tax_silva'
additions = [simple_rec('AAACTGGGGCTAATCCCCCATAGGCCTGGGGTACTGGAAGGTCCCCAGGCCGAAAGGG------GACCGTA-----AGGTCCCGCCCGAGGATGGGCCGGCGGCCGATTAGGTAGTTGGTGGGGTAACGGCCCACCAAG--CCGAAGATCGGTACGGGCC-GTGAGAGCGGGAGCCCGGAGATGGACA---CTGAGACACGGGTCCAGGCCCTACGGGGCGCAGCAGGCGCGAAACC-TCCGCAATGCGGGAAACCGCGACGGGGGGACCCCCAGTGCCGTGCCTCTGGC-----ACGGCTTTTCCGGAGTG-TAAAAAGCTCCGGGAATAAGGGCTGGGCAAGGCCGGTGGC-AGCCGCCGCGGTAATACCGGCGGCCCGAGTGGTGGCCACTATTATTGGGCCTAAAGCGGCCGTAGCCGGGCCCGTAAGTCCCTGGCG-AAATCCCACGGCTCAACCGTGGGGCTCGCTGGGGATACTGCGG-GCCTTGGGACCGGGAGAGGCCGGGGGTACC-CCCGGGGTAGGGGTGAAATCCTATAATCCCGGGGGGACCGCCAGT-GGCGAAGGCGCCC--GGCTGGAACGGGTCCGACGGTGAGGGCCGAAGGCC-AGGGGAGCGAACCGGATTAGATACCCGGGTAGTCCTGGCTGTAAAGGATGCGGGCTAGGTGTCGGGCGAG-CTTCGAGCTCGC-CCGGTGCCGTAGGGAAGCCGTTAAGCCCGCCGCC-TGGGGAGTACGGCCGCAAGGCT-GAAACTTAAAGGAATT-GGCGGGGGAGC-ACTACAAGGGGTGGAGCGTGCGGTTTAATTGGATTCAACGCCGGGAACCTCACCGGGGGCGACGGCAGGATGAA-GGCCAGGCTGAAGGTCTTGCCGGACGCGCCGAGAGGAG-----------------------------------GTGCATGGCCGCCGTCAGCTCGTACCGTGAGGCGTCCA-CTTAAGTGTGGTAACGAGCGAGACCCGC--GCCCCCAGTTGCCAGTCCCTCCCGCTGGGA---GGGAGGC-ACTCTGGGGGG-ACTGCCGGCGAT-AAGCCGGAGGAAGGGGCGGGCGACGGTAGGTCAGTATG-CCCCGAAACCC-CCGGGCT-ACACGCGCGCTACAATGGGCGGGACAATGGGA-CCCGACCCCGAAAGGGGAAGGGAATCCCCTAAACCCGCCCTCAGTTCGGATCGCGGGCTG-CAACTCGCCCGCGTGAAGC-TGGAAT-CCCTAGTACCCGCGCGTCATCATCGCGCGGCGAATACGTCCCTGCTCCTTGCACACACCGCCCGTCACTCCACCCGAG-CGGGGCCC-GGGTGAGGCCCGATCTCCTTCGGGAGGTCGGGTCGAGCCTGGGCTC-CGTGAGGGGGG-AGAAGTCGTAACAAGGTAGCC------------------------------'.replace('-', ''),
'Thermococcus_chitonophagus'),
simple_rec('AAACTGGGATTAATACCCACTAAATGATAATACCTGGAATGGCTTATCATTGAAAGAC-TCTGGAAACATGCTTC-CAGCGTCGCCCAAGG-------------------------------------------------------------------------------GGAGCCCGGAGATGGAAA---CTGAGACAAGGTTCCAGGCCCTACGGGGCGCAGCAGGCGCGAAACC-TCCACAATGCGCGAAAGCGTGATGGGGTTATCCCGAGTGCCGTCCGATGAGG-----ATGGCTTTTCCTCGGTG-TAAGGATCCGAGGGAATAAAGGGGGGGCAAGACTGGTGTC-AGCCGCCGCGGTAATACCAGCTCCCTGAGTGGTAAGGACGATTATTTGGCCTAAAGCGTCCGTAGCCGGCTTATCAAGTCTCTTGTT-AAACCCAGTGATTCAATCATTGACCT-GCAAGAGATACTGTTA-TGCTAGAGGACGGGAGAGGTCGACGG---------GGGTAGGGGTGAAATCCTATAATCCTTGGAGGACCACCAGT-GGCGAAGGCGGTC--GACTAGAACGTGCCTGACGGTGAGGGACGAAAGCT-GGGGGAGCGAACCGGATTAGATACCCGGGTAGTCCCAGCTGTAAACGATGCGGGCTAGGTGTTGGGGTAG-CTACGAGCTACT-CCAGTGCCGCAGAGAAGTTGTTAAGCCCGCCGCC-TGGGGAGTACGGCCGCAAGGCT-GAAACTTAAAGGAATT-GGCGGGGGAGC-ACCACAAGGGGTGAAGGCTGCGGTTTAATTGGAGTCAACGCCGGGAACCTTACCGGGGCTGACAGCAGAGTGAA-GGCCAGACTGAAGATCTTGCCAGACAAGCTGAGAGGAGGTGCATGAAGATCTTGCCAGACAAGCTGAGAGGAGGTGCATGGCCGTCGCCAGTTCGTGCCGTGAGGTGTCCT-GTTAAGTCAGGCAACGAACGAGACCCCC--ACTGTTAGTTGCCAGCGAATTCCAACGGAAT--GTCGGGC-ACACTAACAGG-ACTGCCACCGAT-AAGGTGGAGGAAGGAGGGGGCAACGGCAGGTCAGTATG-CCCC--------------------------------------------------------------------------------------------------------------GAACTCGCCCTCATGAACA-TGGAAT-CCCTAGTAACCGCGTGTCATCATCGCGCGGTGAATACGTCCCCGCTCCTTGCACACACCGCCCGTCGCTCCATCCAAG-TCGGGTCT-AGATGAGGCGCAGTCTTCT-----TGGCTACGTCGAATCTGGGTTC-GGTGAGGGGGG-AGAAGTCGTAACAAGGTGGCCGTAGGGGAACCTGCGGCCGGATCACCTCCT'.replace('-', ''),
'SMTZ1-55'),
simple_rec('ACTCCGGTTGATCCTGCCGGACCCCACTGCTATCGGGGTAGGACTTAACCATGCGAGTTGTGCGTCCCCAAGCCATGGTGGGGGCGCGGCATACGGCTCAGTAACACGTGGCTAACCTAGCCTTTGGACGGGGACAACCCCGGGAAACTGGGGCTAATCCCCGATGGGTGGGAAGGCCTGGAATGGTTTCCCACCGAAAGGGCGTCTGAACCATGCTTCAGGCGTTGCCGAAGGATGGGGCCGCGGCCGATCAGGTTGTTGGTGAGGTAACGGCTCACCAAGCCTATAACCGGTACGGGCCGTGAGAGCGGGAGCCCGGAGATGGGCACTGAGACAAGGGCCCAGGCCCTACGGGGCGCAGCAGGTGCGAAAACTCCGCGATGCGCGAAAGCGTGACGGGGCTATCCCGAGTGCCGTCCGCTGAGGATGGCTTTTCCCCGGTGTAGGGAGCCGGGGGAATAAGGAGAGGGCAAGTCTGGTGTCAGCCGCCGCGGTAATACCAGCTCTCCGAGTGGTGGGGACAATTATTGGGCTTAAAGCGTCCGTAGCCGGCCCATCAAGTCTCTTGTTAAATCCAGCGATCCAATCGCTGGACTGCGGGAGATACTGCTGGGCTAGGGGGCGGGAGAAGCCGATGGTATTCTCGGGGTAGGGGTGAAATCCTATAATCCCGGGAGGACCACCAGTGGCGTAGGCGGTCGGCTAGAACGCGCCCGACGGTGAGGGACGAAAGCTGGGGGAGCGAACCGGATTAGATACCCGGGTAGTCCCAGCCGTAAACGATGCGGGCTAGGTGTTGGGGTGGCTACGAGCCACCCCAGTGCCGCATGGAAGCAATTAAGCCCGCCGCCTGGGGAGTACGGCCGCAAGGCTGAAACTTAAAGGAATTGGCGGGGGAGCACCACAAGGGGTGAAGCTTGCGGTTTAATTGGAGTCAACGCCGGGAAAGGAACAGCGTTTTGTTGTTCCTCTGGATACCTTACCGGGGGCGACAGCAGGATGAAGGCCAGATTGAAGGTCTTGCTGGACGAGCTGAGAGGAGGTGCATGGCCGTCGCCAGTTCGTGCCGTGAGGTGTCCTGTTAAGTCAGGTAACGATCGAGACCCACACCCCCAGTTGCTACCTCTTCGGAGGGCACTCTAGGGGTACTGCCGCCGATAAGGCGGAGGAAGGAGTGGGCCACGGCAGGTCAGTATGCCCCGAATCCCCCGGGCCACACGCGAGCTGCAATGGCAAGGACAATGGGTTCTGACCCCGAGAGGGGAAGGTAATCCCGAAACCCTGCCTCAGTTGGGATCGAGGGCTGAAACCCGCCCTCGTGAACATGGAATCCCTAGTAATCGCGGGTCACCAGCCCGCGGTGAATACGTCCCTGCTCCTTGCACACACCGCCCGTCGCTCCATCCGAGTGGGGTTTAGGTGAGGCGTGGTCCTTGTGGCTGTGTCGAATCTAGGCTTCGCGAGGAGGGAGAAGTCGTAACAAGGTGGCCGTAGGGGAACCTGCGGCCGGATCACCTC',
'BA1-16S')
]
#prepare filter
filt = BlastFilter(lambda a: a.hsps[0].align_length > 1100)
filt.AND = BlastFilter(lambda a: all(hsp.score > 500 for hsp in a.hsps))
filt.AND.AND = BlastFilter(lambda a: all(hsp.identities/float(hsp.align_length) > 0.8 for hsp in a.hsps))
#make ring-blast
blast = BlastCLI(self.abort_event)
orig_seqs = blast.ring_blast(query, suns_db, 100, filt, 3)
if not orig_seqs:
print 'No blast results.'
return 1
nseqs = len(orig_seqs)
print 'RingBlast to:\n%s\nreturned %d sequences.\n' % (suns_db, nseqs)
#save an initial alignment
self.fix_ids(orig_seqs)
alifile = '/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.aln.fasta'
with user_message('Aligning retrieved sequences...', '\n'):
if not AlignmentUtils.align(orig_seqs+[query]+additions, outfile=alifile): return 3
#search for additional homologs
add_seqs = blast.ring_blast(orig_seqs, silva_db, 100, filt, 0)
if add_seqs:
self.fix_ids(add_seqs)
print 'RingBlast to:\n%s\nreturned %d additional sequences.\n' % (silva_db, len(add_seqs))
#build an alignment
seqs = orig_seqs+add_seqs+[query]+additions
alifile = '/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.big.aln.fasta'
with user_message('Aligning retrieved sequences...', '\n'):
if not AlignmentUtils.align(seqs, outfile=alifile): return 3
#build a tree
treefile = '/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.big.aln.tre'
if not PhyloUtils.build_fast_tree(alifile, treefile): return 4
#annotate the tree
with open('/home/allis/Documents/INMI/16S/SSBaF4-SSBaR4-1_243072232-iPCR-report.txt') as inp:
# SSBaF4-SSBaR4_65397396-iPCR-report.txt
sids = set()
len_re = re.compile(r'(\s|^)(\d+)(\sbp|\\s*:)?', re.MULTILINE)
entry = False
cur_sid = None
cur_len = -1
for l in inp:
if l == '========= histograms and electrophorograms of PCR products of each hit =========': break
if l.startswith('---'):
entry = False
if cur_sid and cur_len > 0 and abs(cur_len-920) < 60:
sids.add(cur_sid)
cur_sid = None
cur_len = -1
continue
if entry or '#' in l:
entry = True
plen = len_re.search(l)
if plen: cur_len = int(plen.group(2))
sid = BlastID.extract(l)[0]
if sid: cur_sid = sid
organisms = Organisms.from_records(seqs)
if PhyloUtils.annotate_tree(treefile, organisms,
reroot_at='Thermococcus_chitonophagus',
# beautify_leafs=True,
# collapse_taxa=['miscellaneous crenarchaeotic group', 'thaumarchaeota'],
# collapse_last=True,
# collapse_hard=True,
mark_leafs=sids,
# [r.id for r in orig_seqs+[query]+additions],
lineage_colors={'miscellaneous crenarchaeotic group':(0, 0, 255),
'thaumarchaeta':(255,0,0)},
top_lineage=Lineage('archaea')): return 0
return 2
def _main(self):
min_prod = 400
silva_db = '/home/allis/Documents/INMI/SILVA-DB/SILVA_123_SSURef_Nr99_tax_silva.fasta'
alifile = '/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.aln.fasta'
add_filename = FilenameParser.strip_ext(
alifile) + '.with_additions.fasta'
outgroups = [
'Thermococcus_chitonophagus', 'SMTZ1-55',
'contig72135_1581_sunspring_meta'
]
add = ['KF836721.1.1270', 'EU635905.1.1323']
exclude = [
] #['Thermococcus_chitonophagus', 'SMTZ1-55', 'BA1-16S', 'contig72135_1581_sunspring_meta']
#load alignment
if os.path.isfile(add_filename):
alifile = add_filename
add_filename = ''
with user_message('Loadding initial alignment...', '\n'):
orig_ali = AlignmentUtils.load_first(alifile)
if not orig_ali: return 1
#load homologs
if add_filename:
with user_message('Loadding additional sequences...', '\n'):
add_seqs = []
db = SeqView()
if db.load(silva_db):
for sid in add:
seq = db.get(sid)
if seq: add_seqs.append(seq)
else: print '%s not found in %s' % (sid, silva_db)
#realign data if needed
if add_seqs:
with user_message('Realigning data...', '\n'):
add_filename = FilenameParser.strip_ext(
alifile) + '.with_additions.fasta'
AlignmentUtils.align(
list(orig_ali) + add_seqs, add_filename)
orig_ali = AlignmentUtils.load_first(add_filename)
if not orig_ali: return 2
#process the alignment
ali = orig_ali.remove(*exclude).trim()
for out in outgroups:
if not ali.index(out):
print '%s not found in the alignment' % out
return 3
ali.sort(key=lambda r: 'zzzzzzzz' if r.id in outgroups else r.id)
AlignmentUtils.save(
ali,
'/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.aln.trimmed.fasta'
)
args = dict(plen=(20, 40),
max_mismatches=8,
min_match_mismatches=1,
first_match_mismatches=1,
first_may_match=1,
AT_first=True,
outgroup=len(outgroups))
fprimers = PrimerFinder.find_discriminating_primers(ali, **args)
rprimers = PrimerFinder.find_discriminating_primers(ali,
reverse=True,
**args)
pairs = PrimerFinder.compile_pairs(fprimers, rprimers, min_prod,
'SSBa')
if not pairs:
print '\nNo suitable primer pairs found'
return 3
PrimerFinder.print_pairs(pairs)
orig_ali = PrimerFinder.add_pairs_to_alignment(pairs, orig_ali)
AlignmentUtils.save(
orig_ali,
'/home/allis/Documents/INMI/SunS-metagenome/Bathy/BA2_SunS_16S.with_primers.aln.fasta'
)
print 'Done'
