bedtools/bedtools_subtract

genomics

intervals

subtract

remove

overlap

filter

exclusion

Description

Remove the portion(s) of genomic intervals that are overlapped by other features.

bedtools subtract removes portions of intervals in file A that overlap with intervals
in file B. By default, only the overlapping portions are removed, leaving the
non-overlapping parts of A intervals. This is essential for genomic analysis tasks
like removing repetitive elements, excluding known variants, or filtering out
unwanted regions from interval datasets.

This tool is commonly used for:

Removing repetitive elements or low-complexity regions from analysis
Excluding known polymorphic sites from variant calling regions
Filtering out blacklisted genomic regions from ChIP-seq peaks
Creating clean interval sets by removing overlapping annotations
Generating non-overlapping genomic windows for analysis
Quality control by removing problematic genomic regions

Type

bash_script

License

MIT

Keywords

genomics

intervals

subtract

remove

overlap

filter

exclusion

Links

Repository

Documentation

Contributors

Launch

Run this component

Run the following command to execute this component with Nextflow:

cat > params.yaml <<'EOM'  
output: "$id.$key.output.bed"  
id: "run"  
publish_dir: "output/"  
EOM

nextflow run https://packages.viash-hub.com/vsh/biobox.git \  
  -revision v0.4.0 \  
  -main-script target/nextflow/bedtools/bedtools_subtract/main.nf \  
  -params-file params.yaml

Inputs

Name	Type & Properties	Description
--input_a -a	file required	Input file A containing genomic intervals to subtract from. Format: BED, GFF, or VCF file with genomic coordinates Content: Intervals from which overlapping portions will be removed Usage: This is the primary input file that will be modified Requirements: Standard genomic interval format Output: Modified intervals with overlapping regions removed
--input_b -b	file required	Input file B containing genomic intervals to subtract. Format: BED, GFF, or VCF file with genomic coordinates Content: Intervals that will be removed from file A Usage: Regions in this file will be subtracted from input_a intervals Requirements: Standard genomic interval format Effect: Any overlap with these intervals will be removed from A

Name

Type & Properties

--input_a

-a

file

required

--input_b

-b

file

required

Outputs

Name	Type & Properties	Description
--output	file required output	Output file with subtracted intervals. Format: Same format as input file A Content: Modified intervals from A with B regions removed Behavior: Only non-overlapping portions of A intervals are retained Special cases: Intervals completely overlapped by B are omitted entirely

Name

Type & Properties

--output

file

required

output

Overlap Options

Name	Type & Properties	Description
--min_overlap_a -f	double	Minimum overlap required as a fraction of A interval. Default: 1E-9 (essentially 1 base pair) Range: 0.0 to 1.0 Usage: B must overlap at least this fraction of A for subtraction Example: 0.5 requires B to overlap at least 50% of A interval
--min_overlap_b -F	double	Minimum overlap required as a fraction of B interval. Default: 1E-9 (essentially 1 base pair) Range: 0.0 to 1.0 Usage: A must overlap at least this fraction of B for subtraction Example: 0.3 requires A to overlap at least 30% of B interval
--reciprocal -r	boolean_true	Require that the fraction overlap be reciprocal for A AND B. Usage: Both A and B must overlap each other by the specified fractions Example: With -f 0.9, B must overlap 90% of A AND A must overlap 90% of B Effect: More stringent overlap requirement than -f or -F alone Default: false (only one-way overlap required)
--either_overlap -e	boolean_true	Require that the minimum fraction be satisfied for A OR B. Usage: Either the -f fraction OR the -F fraction must be satisfied Example: With -f 0.9 and -F 0.1, either 90% of A OR 10% of B must be covered Default: false (both fractions must be satisfied if both specified)

Name

Type & Properties

--min_overlap_a

-f

double

--min_overlap_b

-F

double

--reciprocal

-r

boolean_true

--either_overlap

-e

boolean_true

Strand Options

Name	Type & Properties	Description
--same_strand -s	boolean_true	Require same strandedness for overlap detection. Usage: Only subtract B intervals that overlap A on the same strand Applications: Strand-specific analysis, gene annotation filtering Default: false (overlaps reported without respect to strand)
--opposite_strand -S	boolean_true	Require different strandedness for overlap detection. Usage: Only subtract B intervals that overlap A on the opposite strand Applications: Antisense filtering, strand-specific exclusions Default: false (overlaps reported without respect to strand)

Name

Type & Properties

--same_strand

-s

boolean_true

--opposite_strand

-S

boolean_true

Subtraction Behavior

Name	Type & Properties	Description
--remove_entire -A	boolean_true	Remove entire feature if any overlap is found. Behavior: Instead of partial subtraction, remove entire A interval if any overlap with B Usage: Binary filtering - intervals are either kept completely or removed completely Applications: Quality filtering, blacklist exclusion Default: false (only overlapping portions are subtracted)
--remove_if_all_overlap -N	boolean_true	Remove entire feature based on sum of all B feature overlaps. Usage: Same as -A but considers cumulative overlap from multiple B features Behavior: Remove A if total overlap from all B features meets threshold Applications: Complex filtering based on multiple overlapping annotations Requires: Used with -f option for fraction threshold

Name

Type & Properties

--remove_entire

-A

boolean_true

--remove_if_all_overlap

-N

boolean_true

Output Format Options

Name	Type & Properties	Description
--write_original_b -wb	boolean_true	Write the original entry in B for each overlap. Output: Each A interval followed by overlapping B intervals Usage: Track which B features caused the subtraction Restrictions: Overlap filtering by -f and -r still applies Applications: Detailed overlap reporting, annotation tracking
--write_overlap_counts -wo	boolean_true	Write original A and B entries plus the number of base pairs of overlap. Output: A interval, B interval, and overlap count in base pairs Usage: Quantify the extent of overlap between features Filtering: Only A features with overlap are reported Applications: Overlap statistics, coverage analysis
--output_bed -bed	boolean_true	Convert BAM input to BED format output. Usage: Only applicable when input is BAM format Effect: Output genomic coordinates in BED format Applications: Converting alignment data to interval format
--include_header -header	boolean_true	Print the header from the A file prior to results. Usage: Preserves metadata and format information from input Applications: Maintaining file structure and annotations Formats: Particularly relevant for VCF and GFF files

Name

Type & Properties

--write_original_b

-wb

boolean_true

--write_overlap_counts

-wo

boolean_true

--output_bed

-bed

boolean_true

--include_header

-header

boolean_true

Performance Options

Name	Type & Properties	Description
--use_split -split	boolean_true	Treat "split" BAM or BED12 entries as distinct BED intervals. Usage: Split complex entries into individual intervals Applications: Detailed analysis of spliced alignments or multi-part features Formats: Applies to BAM alignments and BED12 format
--sorted_input -sorted	boolean_true	Use the "chromsweep" algorithm for sorted input. Requirements: Input must be sorted by chromosome then start coordinate Performance: Much faster for large sorted files Usage: Optimize processing for pre-sorted genomic data Sort command: sort -k1,1 -k2,2n input.bed
--genome_file -g	file	Genome file to enforce consistent chromosome sort order. Format: Tab-delimited file with chromosome names and lengths Usage: Only applies when used with -sorted option Purpose: Ensure consistent chromosome ordering across input files Example: chr1\t249250621
--no_name_check -nonamecheck	boolean_true	Don't throw error for different chromosome naming conventions. Usage: For sorted data with mixed naming (e.g., "chr1" vs "chr01") Applications: Working with data from different sources Safety: Allows processing despite naming inconsistencies
--no_buffer -nobuf	boolean_true	Disable output buffering for real-time processing. Effect: Each line printed immediately instead of buffered Trade-off: Slower output but enables real-time processing Applications: Pipeline integration, streaming analysis
--input_buffer -iobuf	string	Amount of memory to allocate for input buffer. Format: Integer with optional K/M/G suffix Examples: "1G", "512M", "2048K" Usage: Larger buffers can improve I/O performance for large files Note: Currently has no effect with compressed files

Name

Type & Properties

--use_split

-split

boolean_true

--sorted_input

-sorted

boolean_true

--genome_file

-g

file

--no_name_check

-nonamecheck

boolean_true

--no_buffer

-nobuf

boolean_true

--input_buffer

-iobuf

string