def _total_alignment_score(self, seq_name):
tmpdir = tempfile.mkdtemp(prefix='tmp.get_total_aln_score.', dir=os.getcwd())
tmp_bam = os.path.join(tmpdir, 'tmp.get_total_alignment_score.bam')
tmp_fa = os.path.join(tmpdir, 'tmp.get_total_alignment_score.ref.fa')
faidx.write_fa_subset(
[seq_name],
self.references_fa,
tmp_fa,
samtools_exe=self.samtools_exe,
verbose=True,
verbose_filehandle=self.log_fh
)
mapping.run_bowtie2(
self.reads1,
self.reads2,
tmp_fa,
tmp_bam[:-4],
threads=self.threads,
samtools=self.samtools_exe,
bowtie2=self.bowtie2_exe,
bowtie2_preset=self.bowtie2_preset,
verbose=True,
verbose_filehandle=self.log_fh
)
score = mapping.get_total_alignment_score(tmp_bam)
shutil.rmtree(tmpdir)
return score
def _get_total_alignment_score(self, gene_name):
tmp_bam = os.path.join(self.root_dir,
'tmp.get_total_alignment_score.bam')
assert not os.path.exists(tmp_bam)
tmp_fa = os.path.join(self.root_dir,
'tmp.get_total_alignment_score.ref.fa')
assert not os.path.exists(tmp_fa)
faidx.write_fa_subset([gene_name],
self.genes_fa,
tmp_fa,
samtools_exe=self.samtools_exe,
verbose=self.verbose)
mapping.run_bowtie2(
self.reads1,
self.reads2,
tmp_fa,
tmp_bam[:-4],
threads=self.threads,
samtools=self.samtools_exe,
bowtie2=self.bowtie2_exe,
bowtie2_preset=self.bowtie2_preset,
verbose=self.verbose,
)
score = mapping.get_total_alignment_score(tmp_bam)
os.unlink(tmp_bam)
os.unlink(tmp_fa)
os.unlink(tmp_fa + '.fai')
return score
def test_write_fa_subset(self):
'''test write_fa_subset'''
infile = os.path.join(data_dir, 'faidx_test_write_fa_subset.in.fa')
expected = os.path.join(data_dir, 'faidx_test_write_fa_subset.out.fa')
tmpfile = 'tmp.test_write_fa_subset.out.fa'
faidx.write_fa_subset(['seq1', 'seq3', 'seq4'], infile, tmpfile)
self.assertTrue(filecmp.cmp(expected, tmpfile, shallow=False))
os.unlink(tmpfile)
def _choose_best_gene(self):
gene_name = self._get_best_gene_by_alignment_score()
if gene_name is None:
return None
faidx.write_fa_subset([gene_name], self.genes_fa, self.gene_fa, samtools_exe=self.samtools_exe, verbose=self.verbose)
seqs = {}
pyfastaq.tasks.file_to_dict(self.gene_fa, seqs)
assert len(seqs) == 1
return list(seqs.values())[0]
def best_seq(self, outfile):
'''Finds the closest matchng sequence, writes it to a FASTA file, and returns it as a pyfastaq.sequences.Fasta object'''
seq_name = self._get_best_seq_by_alignment_score()
if seq_name is None:
return None
faidx.write_fa_subset([seq_name], self.references_fa, outfile, samtools_exe=self.samtools_exe, verbose=True, verbose_filehandle=self.log_fh)
seqs = {}
pyfastaq.tasks.file_to_dict(outfile, seqs)
assert len(seqs) == 1
return list(seqs.values())[0]
def _choose_best_gene(self):
gene_name = self._get_best_gene_by_alignment_score()
if gene_name is None:
return None
faidx.write_fa_subset([gene_name],
self.genes_fa,
self.gene_fa,
samtools_exe=self.samtools_exe,
verbose=self.verbose)
seqs = {}
pyfastaq.tasks.file_to_dict(self.gene_fa, seqs)
assert len(seqs) == 1
return list(seqs.values())[0]
def _init_and_run_clusters(self):
if len(self.cluster_to_dir) == 0:
raise Error('Did not get any reads mapped to genes. Cannot continue')
counter = 0
for gene in sorted(self.cluster_to_dir):
counter += 1
if self.verbose:
print('\nAssembling cluster', counter, 'of', str(len(self.cluster_to_dir)))
new_dir = self.cluster_to_dir[gene]
faidx.write_fa_subset(
self.cluster_ids[gene],
self.db_fasta,
os.path.join(new_dir, 'genes.fa'),
samtools_exe=self.samtools_exe,
verbose=self.verbose
)
self.clusters[gene] = cluster.Cluster(
new_dir,
gene,
assembly_kmer=self.assembly_kmer,
assembler=self.assembler,
max_insert=self.insert_proper_pair_max,
min_scaff_depth=self.min_scaff_depth,
nucmer_min_id=self.nucmer_min_id,
nucmer_min_len=self.nucmer_min_len,
nucmer_breaklen=self.nucmer_breaklen,
sspace_k=self.min_scaff_depth,
reads_insert=self.insert_size,
sspace_sd=self.insert_sspace_sd,
threads=self.threads,
assembled_threshold=self.assembled_threshold,
unique_threshold=self.unique_threshold,
verbose=self.verbose,
bcftools_exe=self.bcftools_exe,
gapfiller_exe=self.gapfiller_exe,
samtools_exe=self.samtools_exe,
bowtie2_exe=self.bowtie2_exe,
bowtie2_preset=self.bowtie2_preset,
spades_exe=self.spades_exe,
sspace_exe=self.sspace_exe,
velvet_exe=self.velvet,
spades_other=self.spades_other,
clean=self.clean,
)
self.clusters[gene].run()
def _init_and_run_clusters(self):
if len(self.cluster_to_dir) == 0:
raise Error('Did not get any reads mapped to genes. Cannot continue')
counter = 0
for gene in sorted(self.cluster_to_dir):
counter += 1
if self.verbose:
print('\nAssembling cluster', counter, 'of', str(len(self.cluster_to_dir)))
new_dir = self.cluster_to_dir[gene]
faidx.write_fa_subset(
self.cluster_ids[gene],
self.db_fasta,
os.path.join(new_dir, 'genes.fa'),
samtools_exe=self.samtools_exe,
verbose=self.verbose
)
self.clusters[gene] = cluster.Cluster(
new_dir,
gene,
assembly_kmer=self.assembly_kmer,
assembler=self.assembler,
max_insert=self.insert_proper_pair_max,
min_scaff_depth=self.min_scaff_depth,
nucmer_min_id=self.nucmer_min_id,
nucmer_min_len=self.nucmer_min_len,
nucmer_breaklen=self.nucmer_breaklen,
sspace_k=self.min_scaff_depth,
reads_insert=self.insert_size,
sspace_sd=self.insert_sspace_sd,
threads=self.threads,
assembled_threshold=self.assembled_threshold,
unique_threshold=self.unique_threshold,
verbose=self.verbose,
bcftools_exe=self.bcftools_exe,
gapfiller_exe=self.gapfiller_exe,
samtools_exe=self.samtools_exe,
bowtie2_exe=self.bowtie2_exe,
bowtie2_preset=self.bowtie2_preset,
spades_exe=self.spades_exe,
sspace_exe=self.sspace_exe,
velvet_exe=self.velvet,
spades_other=self.spades_other,
clean=self.clean,
)
self.clusters[gene].run()
def _get_total_alignment_score(self, gene_name):
tmp_bam = os.path.join(self.root_dir, 'tmp.get_total_alignment_score.bam')
assert not os.path.exists(tmp_bam)
tmp_fa = os.path.join(self.root_dir, 'tmp.get_total_alignment_score.ref.fa')
assert not os.path.exists(tmp_fa)
faidx.write_fa_subset([gene_name], self.genes_fa, tmp_fa, samtools_exe=self.samtools_exe, verbose=self.verbose)
mapping.run_bowtie2(
self.reads1,
self.reads2,
tmp_fa,
tmp_bam[:-4],
threads=self.threads,
samtools=self.samtools_exe,
bowtie2=self.bowtie2_exe,
bowtie2_preset=self.bowtie2_preset,
verbose=self.verbose,
)
score = mapping.get_total_alignment_score(tmp_bam)
os.unlink(tmp_bam)
os.unlink(tmp_fa)
os.unlink(tmp_fa + '.fai')
return score
def run(self):
self._assemble_with_fermilite()
self.sequences = {}
# double-check we got some contigs
number_of_contigs = pyfastaq.tasks.count_sequences(self.assembly_contigs) if os.path.exists(self.assembly_contigs) else 0
if number_of_contigs == 0:
self.assembled_ok = False
# This is to make this object picklable, to keep multithreading happy
self.log_fh = None
return
else:
self.assembled_ok = True
if self.assembled_ok:
self._scaffold_with_sspace()
self._gap_fill_with_gapfiller()
pyfastaq.tasks.filter(self.gapfilled_scaffolds, self.gapfilled_length_filtered, minlength=self.min_scaff_length)
if pyfastaq.tasks.count_sequences(self.gapfilled_length_filtered) == 0:
self.assembled_ok = False
# This is to make this object picklable, to keep multithreading happy
self.log_fh = None
return
masher = mash.Masher(self.ref_fastas, self.gapfilled_length_filtered, self.log_fh, self.extern_progs)
self.ref_seq_name = masher.run(self.mash_dist_file)
if self.ref_seq_name is None:
print('Could not determine closest reference sequence', file=self.log_fh)
self.log_fh = None
return
faidx.write_fa_subset({self.ref_seq_name}, self.ref_fastas, self.ref_fasta, samtools_exe=self.extern_progs.exe('samtools'), verbose=True, verbose_filehandle=self.log_fh)
print('Closest reference sequence according to mash: ', self.ref_seq_name, file=self.log_fh)
contigs_both_strands = self._fix_contig_orientation(self.gapfilled_length_filtered, self.ref_fasta, self.final_assembly_fa, min_id=self.nucmer_min_id, min_length=self.nucmer_min_len, breaklen=self.nucmer_breaklen)
self.has_contigs_on_both_strands = len(contigs_both_strands) > 0
pyfastaq.tasks.file_to_dict(self.final_assembly_fa, self.sequences)
mapping.run_bowtie2(
self.reads1,
self.reads2,
self.final_assembly_fa,
self.final_assembly_bam[:-4],
threads=1,
sort=True,
samtools=self.extern_progs.exe('samtools'),
bowtie2=self.extern_progs.exe('bowtie2'),
verbose=True,
verbose_filehandle=self.log_fh
)
self.scaff_graph_ok = self._parse_bam(self.sequences, self.final_assembly_bam, self.min_scaff_depth, self.max_insert)
print('Scaffolding graph is OK:', self.scaff_graph_ok, file=self.log_fh)
if self.clean:
for suffix in ['soft_clipped', 'unmapped_mates', 'scaff']:
filename = self.final_assembly_bam + '.' + suffix
print('Deleting file', filename, file=self.log_fh)
os.unlink(filename)
# This is to make this object picklable, to keep multithreading happy
self.log_fh = None
