BFBArchitect

BFBArchitect is a tool for detecting and reconstructing breakage-fusion-bridge (BFB) cycles from long-read sequencing (currently support Oxford Nanopore).

Prerequisites

• CNVkit>=0.9.10 (https://cnvkit.readthedocs.io/en/master/quickstart.html)
• pandas>=2.3.3 (https://pandas.pydata.org/docs/whatsnew/index.html)
• PuLP>=3.3.0 (https://coin-or.github.io/pulp/main/includeme.html)
• pysam>=0.23.3 (https://pysam.readthedocs.io/en/latest/release.html)
• Python>=3.12.8 (https://www.python.org/downloads/release/python-3128/)

Installation

BFBArchitect can be installed and run on most modern Unix-like operating systems (e.g. Ubuntu 18.04+, CentOS 7+, macOS). It requires python>=3.8 and the above dependencies. Please follow the instructions to install (more installation options will be provided soon):

1. Pull the source code

   git clone git@github.com:AmpliconSuite/BFBArchitect.git
   cd /path/to/BFBArchitect
   

2. Create a virtual environment (optional)

   python3 -m venv BFBArchitect_venv
   source BFBArchitect_venv/bin/activate
   

3. Install dependencies locally

   pip install .
   

Running

Before running BFBArchitect, genome-wide copy number (CN) calls must be generated from the aligned long-read data by running the follow script:

/path/to/BFBArchitect/scripts/call_cnvs.sh <input.bam> /path/to/BFBArchitect/scripts/hg38full_ref_5k.cnn <output_dir>

This will create a file called [input].cns, which is a required argument in BFBArchitect.

Then run BFBArchitect to reconstruct potential BFB sequences for any genomic region chrom:start-end with copy number amplification. (The amplicon region can be detected by standard pipelines like CoRAL.)

Usage

python /path/to/BFBArchitect.py --bam <input.bam> --cns <input.cns> --regions <chrom:start-end> --output_prefix <dir/output_prefix> [--segmentation] [--deletion] [--coverage <sequencing coverage>]

Required arguments

• --bam <.bam file>: Aligned long reads
• --cns <.cns file>: The .cns file from genome-wide copy number calling
• --regions : A string that represents the amplified genomic region (e.g. chr1:1-1000000)
• --output_prefix : The directory and prefix for all output files

Optional arguments

• --segmentation: Consider copy number variation when segmenting the amplicon region.
• --deletion: Handle deletion when reconstructing BFB sequences.
• --coverage : Sequencing coverage (if provided, estimating coverage from cns will be skipped)

Output

• graph.txt: A text file describing the segment and structural variant information of a breakpoint graph constructed from the amplicon region.
• cycles.txt: A text file including the reconstructed BFB sequences.
• reads.txt: A text file storing information of supporting reads for structural variants.

Sample run

Please download the sample input from this link, which includes sample.sorted.bam, sample.sorted.cns, and sample.sorted.cnr (for visualization). Run the following command:

python /path/to/BFBArchitect/src/BFBArchitect.py --bam BFBArchitect_input/sample.sorted.bam --cns BFBArchitect_input/sample.sorted.cns --regions chr7:120000000-125000000 --output_prefix sample --coverage 15.0

The following files will be output:

1. sample_graph.txt

    SequenceEdge: StartPosition, EndPosition, PredictedCN, AverageCoverage, Size, NumberOfLongReads
    sequence	chr7:120000001-	chr7:123649137+	1	15.953404599498457	3649137	11764
    sequence	chr7:123649138-	chr7:124186212+	10	84.07632453567938	537075	8843
    sequence	chr7:124186213-	chr7:124347510+	5	48.4874021996553	161298	1607
    sequence	chr7:124347511-	chr7:124740114+	3	31.59089311367179	392604	2494
    sequence	chr7:124740115-	chr7:124999999+	7	61.75353714142794	259885	3388
    BreakpointEdge: StartPosition->EndPosition, PredictedCN, NumberOfLongReads
    concordant	chr7:123649137+->chr7:123649138-	1	50
    concordant	chr7:124186212+->chr7:124186213-	5	66
    concordant	chr7:124347510+->chr7:124347511-	3	40
    concordant	chr7:124740114+->chr7:124740115-	3	46
    discordant	chr7:123649138-->chr7:123650665-	4	33
    discordant	chr7:124184685+->chr7:124186212+	2	14
    discordant	chr7:124346947+->chr7:124347510+	1	7
    discordant	chr7:124740115-->chr7:124740275-	2	17
    discordant	chr7:124807844+->chr7:124840986-	7	51
    discordant	chr7:124998462+->chr7:124999999+	4	29
   

2. sample_cycles.txt

    Interval	1	chr7	120000001	124999999
    List of cycle segments
    Segment	1	chr7	120000001	123649137
    Segment	2	chr7	123649138	124186212
    Segment	3	chr7	124186213	124347510
    Segment	4	chr7	124347511	124740114
    Segment	5	chr7	124740115	124999999
    List of longest subpath constraints
    Path=1;Copy_count=1;Segments=1+,2+,3+,4+,5+,5-,5+,5-,5+,5-,4-,3-,2-,2+,2-,2+,3+,3-,2-,2+,2-,2+,3+,4+,5+,5-;Path_constraints_satisfied=;Score=0.30515965787039095;Multiplicity=1
   

3. sample_reads.txt

    SV	SV_type	TST	#Support_reads	Query_gaps	Support_reads
    ...
   

After reconstructing BFB sequences, visualization can be generated by running the following command:

python ~/BFBArchitect/src/BFBVisualizer.py --graph sample_graph.txt --cycle sample_cycles.txt --cnr BFBArchitect_input/sample.sorted.cnr --output_prefix sample

Contact

BFBArchitect is developed and maintained by Bafna Lab at UC San Diego. Please raise an issue or reach out to chl221@ucsd.edu if you have any questions.

License

This project is is licensed under the BSD 3-Clause License - see the LICENSE.txt file for details