diff --git a/fgpyo/sam/__init__.py b/fgpyo/sam/__init__.py index 454ed12..99cbea8 100644 --- a/fgpyo/sam/__init__.py +++ b/fgpyo/sam/__init__.py @@ -162,6 +162,7 @@ from pathlib import Path from typing import IO from typing import Any +from typing import Callable from typing import Dict from typing import Iterable from typing import Iterator @@ -176,6 +177,7 @@ from pysam import AlignedSegment from pysam import AlignmentFile as SamFile from pysam import AlignmentHeader as SamHeader +from typing_extensions import deprecated import fgpyo.io from fgpyo.collections import PeekableIterator @@ -802,38 +804,84 @@ def isize(r1: AlignedSegment, r2: AlignedSegment) -> int: return r2_pos - r1_pos -def set_pair_info(r1: AlignedSegment, r2: AlignedSegment, proper_pair: bool = True) -> None: - """Resets mate pair information between reads in a pair. Requires that both r1 - and r2 are mapped. Can be handed reads that already have pairing flags setup or - independent R1 and R2 records that are currently flagged as SE reads. +def set_mate_info( + r1: AlignedSegment, + r2: AlignedSegment, + is_proper_pair: Callable[[AlignedSegment, AlignedSegment], bool] = properly_paired, +) -> None: + """Resets mate pair information between reads in a pair. + + Args: + r1: Read 1 (first read in the template). + r2: Read 2 with the same query name as r1 (second read in the template). + is_proper_pair: A function that takes the two reads and determines proper pair status. + """ + if r1.query_name != r2.query_name: + raise ValueError("Cannot pair reads with different query names!") + + for src, dest in [(r1, r2), (r2, r1)]: + dest.next_reference_id = src.reference_id + dest.next_reference_name = src.reference_name + dest.next_reference_start = src.reference_start + dest.mate_is_forward = src.is_forward + dest.mate_is_mapped = src.is_mapped + dest.set_tag("MC", src.cigarstring) + dest.set_tag("MQ", src.mapping_quality) + + template_length = isize(r1, r2) + r1.template_length = template_length + r2.template_length = -template_length + + proper_pair = is_proper_pair(r1, r2) + r1.is_proper_pair = proper_pair + r2.is_proper_pair = proper_pair + + +def set_as_pairs( + r1: AlignedSegment, + r2: AlignedSegment, + is_proper_pair: Callable[[AlignedSegment, AlignedSegment], bool] = properly_paired, +) -> None: + """Forces the two reads to become pairs as long as they share the same query name. + + This function will take two reads, as long as they have the same query name, and make them + pairs. First, the reads are marked as pairs and then first and second read flags are set. + Finally, all mate information is reset upon both reads. + + This function is useful for taking once-single-end reads and making them pairs. Args: - r1: read 1 - r2: read 2 with the same queryname as r1 + r1: Read 1 (first read in the template). + r2: Read 2 with the same query name as r1 (second read in the template). + is_proper_pair: A function that takes the two reads and determines proper pair status. """ if r1.query_name != r2.query_name: - raise ValueError("Cannot set pair info on reads with different query names!") + raise ValueError("Cannot pair reads with different query names!") - for r in [r1, r2]: + for r in (r1, r2): r.is_paired = True - r.is_proper_pair = proper_pair r1.is_read1 = True r1.is_read2 = False r2.is_read2 = True r2.is_read1 = False - for src, dest in [(r1, r2), (r2, r1)]: - dest.next_reference_id = src.reference_id - dest.next_reference_start = src.reference_start - dest.mate_is_reverse = src.is_reverse - dest.mate_is_unmapped = src.mate_is_unmapped - dest.set_tag("MC", src.cigarstring) - dest.set_tag("MQ", src.mapping_quality) + set_mate_info(r1=r1, r2=r2, is_proper_pair=is_proper_pair) + + +@deprecated("Use `set_as_pairs()` instead. Deprecated as of fgpyo 0.8.0.") +def set_pair_info(r1: AlignedSegment, r2: AlignedSegment, proper_pair: bool = True) -> None: + """Resets mate pair information between reads in a pair. - insert_size = isize(r1, r2) - r1.template_length = insert_size - r2.template_length = -insert_size + Requires that both r1 and r2 are mapped. Can be handed reads that already have pairing flags + setup or independent R1 and R2 records that are currently flagged as SE reads. + + Args: + r1: Read 1 (first read in the template). + r2: Read 2 with the same query name as r1 (second read in the template). + proper_pair: whether the pair is proper or not. + """ + set_as_pairs(r1, r2, lambda r1, r2: proper_pair) @attr.s(frozen=True, auto_attribs=True) diff --git a/fgpyo/sam/builder.py b/fgpyo/sam/builder.py index 4a7b58d..53997c2 100755 --- a/fgpyo/sam/builder.py +++ b/fgpyo/sam/builder.py @@ -264,70 +264,6 @@ def _set_length_dependent_fields( if not rec.is_unmapped: rec.cigarstring = cigar if cigar else f"{length}M" - def _set_mate_info(self, r1: pysam.AlignedSegment, r2: pysam.AlignedSegment) -> None: - """Sets the mate information on a pair of sam records. - - Handles cases where both reads are mapped, one of the two reads is unmapped or both reads - are unmapped. - - Args: - r1: the first read in the pair - r2: the sceond read in the pair - """ - for rec in r1, r2: - rec.template_length = 0 - rec.is_proper_pair = False - - if r1.is_unmapped and r2.is_unmapped: - # If they're both unmapped just clean the records up - for rec, other in [(r1, r2), (r2, r1)]: - rec.reference_id = sam.NO_REF_INDEX - rec.next_reference_id = sam.NO_REF_INDEX - rec.reference_start = sam.NO_REF_POS - rec.next_reference_start = sam.NO_REF_POS - rec.is_unmapped = True - rec.mate_is_unmapped = True - rec.is_proper_pair = False - rec.mate_is_reverse = other.is_reverse - - elif r1.is_unmapped or r2.is_unmapped: - # If only one is mapped/unmapped copy over the relevant stuff - (m, u) = (r1, r2) if r2.is_unmapped else (r2, r1) - u.reference_id = m.reference_id - u.reference_start = m.reference_start - u.next_reference_id = m.reference_id - u.next_reference_start = m.reference_start - u.mate_is_reverse = m.is_reverse - u.mate_is_unmapped = False - u.set_tag("MC", m.cigarstring) - - m.next_reference_id = u.reference_id - m.next_reference_start = u.reference_start - m.mate_is_reverse = u.is_reverse - m.mate_is_unmapped = True - - else: - # Else they are both mapped - for rec, other in [(r1, r2), (r2, r1)]: - rec.next_reference_id = other.reference_id - rec.next_reference_start = other.reference_start - rec.mate_is_reverse = other.is_reverse - rec.mate_is_unmapped = False - rec.set_tag("MC", other.cigarstring) - - if r1.reference_id == r2.reference_id: - r1p = r1.reference_end if r1.is_reverse else r1.reference_start - r2p = r2.reference_end if r2.is_reverse else r2.reference_start - r1.template_length = r2p - r1p - r2.template_length = r1p - r2p - - # Arbitrarily set proper pair if the we have an FR pair with isize <= 1000 - if r1.is_reverse != r2.is_reverse and abs(r1.template_length) <= 1000: - fpos, rpos = (r2p, r1p) if r1.is_reverse else (r1p, r2p) - if fpos < rpos: - r1.is_proper_pair = True - r2.is_proper_pair = True - def rg(self) -> Dict[str, Any]: """Returns the single read group that is defined in the header.""" # The `RG` field contains a list of read group mappings @@ -444,8 +380,16 @@ def add_pair( raise ValueError("Cannot use chrom in combination with chrom1 or chrom2") chrom = sam.NO_REF_NAME if chrom is None else chrom - chrom1 = next(c for c in (chrom1, chrom) if c is not None) - chrom2 = next(c for c in (chrom2, chrom) if c is not None) + + if start1 != sam.NO_REF_POS: + chrom1 = next(c for c in (chrom1, chrom) if c is not None) + else: + chrom1 = sam.NO_REF_NAME + + if start2 != sam.NO_REF_POS: + chrom2 = next(c for c in (chrom2, chrom) if c is not None) + else: + chrom2 = sam.NO_REF_NAME if chrom1 == sam.NO_REF_NAME and start1 != sam.NO_REF_POS: raise ValueError("start1 cannot be used on its own - specify chrom or chrom1") @@ -468,7 +412,7 @@ def add_pair( ) # Sync up mate info and we're done! 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a/pyproject.toml +++ b/pyproject.toml @@ -26,7 +26,7 @@ include = ["LICENSE"] python = ">=3.9.0,<4.0" attrs = ">=19.3.0" pysam = ">=0.22.1" -typing_extensions = { version = ">=3.7.4", python = "<3.12" } +typing_extensions = { version = ">=4.12.2", python = "<3.12" } numpy = [ {version = "^1.26.4", python = ">=3.12"}, {version = "^1.25.2", python = ">=3.9,<3.12"}, diff --git a/tests/fgpyo/sam/test_builder.py b/tests/fgpyo/sam/test_builder.py index f3bbcd5..2b5d795 100755 --- a/tests/fgpyo/sam/test_builder.py +++ b/tests/fgpyo/sam/test_builder.py @@ -114,33 +114,56 @@ def test_unmapped_reads() -> None: assert r1.mate_is_unmapped assert r2.is_unmapped assert not r2.mate_is_unmapped - for rec in r1, r2: - assert rec.reference_name == "chr1" - assert rec.reference_start == 1000 - assert rec.next_reference_name == "chr1" - assert rec.next_reference_start == 1000 + assert r1.reference_name == "chr1" + assert r1.reference_start == 1000 + assert r1.next_reference_name is None + assert r1.next_reference_start == sam.NO_REF_POS + assert r2.reference_name is None + assert r2.reference_start == sam.NO_REF_POS + assert r2.next_reference_name == "chr1" + assert r2.next_reference_start == 1000 r1, r2 = builder.add_pair(chrom="chr1", start2=2000) assert r1.is_unmapped assert not r1.mate_is_unmapped assert not r2.is_unmapped assert r2.mate_is_unmapped - for rec in r1, r2: - assert rec.reference_name == "chr1" - assert rec.reference_start == 2000 - assert rec.next_reference_name == "chr1" - assert rec.next_reference_start == 2000 + assert r1.reference_name is None + assert r1.reference_start == sam.NO_REF_POS + assert r1.next_reference_name == "chr1" + assert r1.next_reference_start == 2000 + assert r2.reference_name == "chr1" + assert r2.reference_start == 2000 + assert r2.next_reference_name is None + assert r2.next_reference_start == sam.NO_REF_POS r1, r2 = builder.add_pair(chrom=sam.NO_REF_NAME) assert r1.is_unmapped assert r1.mate_is_unmapped assert r2.is_unmapped assert r2.mate_is_unmapped - for rec in r1, r2: - assert rec.reference_name is None - assert rec.reference_start == sam.NO_REF_POS - assert rec.next_reference_name is None - assert rec.next_reference_start == sam.NO_REF_POS + assert r1.reference_name is None + assert r1.reference_start == sam.NO_REF_POS + assert r1.next_reference_name is None + assert r1.next_reference_start == sam.NO_REF_POS + assert r2.reference_name is None + assert r2.reference_start == sam.NO_REF_POS + assert r2.next_reference_name is None + assert r2.next_reference_start == sam.NO_REF_POS + + r1, r2 = builder.add_pair(chrom=None) + assert r1.is_unmapped + assert r1.mate_is_unmapped + assert r2.is_unmapped + assert r2.mate_is_unmapped + assert r1.reference_name is None + assert r1.reference_start == sam.NO_REF_POS + assert r1.next_reference_name is None + assert r1.next_reference_start == sam.NO_REF_POS + assert r2.reference_name is None + assert r2.reference_start == sam.NO_REF_POS + assert r2.next_reference_name is None + assert r2.next_reference_start == sam.NO_REF_POS def test_invalid_strand() -> None: diff --git a/tests/fgpyo/sam/test_sam.py b/tests/fgpyo/sam/test_sam.py index 497a35b..fe1f844 100755 --- a/tests/fgpyo/sam/test_sam.py +++ b/tests/fgpyo/sam/test_sam.py @@ -311,19 +311,19 @@ def test_pair_orientation_build_with_r2() -> None: r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = False r2.is_forward = True - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert PairOrientation.build(r1, r2) is PairOrientation.RF r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = True r2.is_forward = True - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert PairOrientation.build(r1, r2) is PairOrientation.TANDEM r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = False r2.is_forward = False - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert PairOrientation.build(r1, r2) is PairOrientation.TANDEM @@ -337,7 +337,7 @@ def test_pair_orientation_is_fr_if_opposite_directions_and_overlapping() -> None r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="10M", start2=100, cigar2="10M") r1.is_reverse = True r2.is_reverse = False - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert PairOrientation.build(r1, r2) is PairOrientation.FR @@ -369,19 +369,19 @@ def test_pair_orientation_build_with_no_r2_but_r2_mapped() -> None: r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = False r2.is_forward = True - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert PairOrientation.build(r1) is PairOrientation.RF r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = True r2.is_forward = True - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert PairOrientation.build(r1) is PairOrientation.TANDEM r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = False r2.is_forward = False - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert PairOrientation.build(r1) is PairOrientation.TANDEM @@ -391,19 +391,19 @@ def test_pair_orientation_build_with_either_unmapped_but_no_r2() -> None: r1, r2 = builder.add_pair() assert r1.is_unmapped assert r2.is_unmapped - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert PairOrientation.build(r1) is None r1, r2 = builder.add_pair(chrom="chr1", start1=100) assert r1.is_mapped assert r2.is_unmapped - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert PairOrientation.build(r1) is None r1, r2 = builder.add_pair(chrom="chr1", start2=100) assert r1.is_unmapped assert r2.is_mapped - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert PairOrientation.build(r1) is None @@ -411,7 +411,7 @@ def test_pair_orientation_build_raises_if_it_cant_find_mate_cigar_tag() -> None: """Test that an exception is raised if we cannot find the mate cigar tag.""" builder = SamBuilder() r1, r2 = builder.add_pair(chrom="chr1", start1=10, start2=30) - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) r1.set_tag("MC", None) # Clear out the MC tag. with pytest.raises(ValueError): @@ -427,7 +427,7 @@ def test_properly_paired_when_actually_proper() -> None: r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="10M", start2=100, cigar2="10M") r1.is_reverse = True r2.is_reverse = False - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert properly_paired(r1, r2) @@ -440,7 +440,7 @@ def test_properly_paired_when_actually_proper_and_no_r2() -> None: r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="10M", start2=100, cigar2="10M") r1.is_reverse = True r2.is_reverse = False - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert properly_paired(r1) @@ -450,19 +450,19 @@ def test_not_properly_paired_if_wrong_orientation() -> None: r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = False r2.is_forward = True - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert not properly_paired(r1, r2) r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = True r2.is_forward = True - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert not properly_paired(r1, r2) r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = False r2.is_forward = False - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert not properly_paired(r1, r2) @@ -472,19 +472,19 @@ def test_not_properly_paired_if_wrong_orientation_and_no_r2() -> None: r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = False r2.is_forward = True - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert not properly_paired(r1) r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = True r2.is_forward = True - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert not properly_paired(r1) r1, r2 = builder.add_pair(chrom="chr1", start1=100, cigar1="115M", start2=250, cigar2="40M") r1.is_forward = False r2.is_forward = False - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert not properly_paired(r1) @@ -492,7 +492,7 @@ def test_not_properly_paired_if_too_far_apart() -> None: """Test that reads are not properly paired if they are too far apart.""" builder = SamBuilder() r1, r2 = builder.add_pair(chrom="chr1", start1=100, start2=100 + 1000) - sam.set_pair_info(r1, r2) + sam.set_mate_info(r1, r2) assert not properly_paired(r1, r2)