Package 'ImprintCapASM'

Run Allele-Specific Methylation Analysis

Description

Performs SNP-phased allele-specific methylation (ASM) analysis across the canonical human imprinted differentially methylated regions (DMRs). Reads are assigned to REF or ALT alleles based on bisulfite-aware SNP detection, and per-allele CpG methylation fractions are computed.

Usage

ASM(
  cpg_snp_file,
  sam_file,
  filter_cpgs_file,
  output_file = NULL,
  sample_type = c("control", "patient"),
  verbose = TRUE
)

Arguments

cpg_snp_file	Character. Path to the Excel output from prepare_cpg_snp_input.
sam_file	Character. Path to the extracted wide BAM file produced by extract_bam_regions.
filter_cpgs_file	Character. Path to the CpG filter/variability reference Excel file (filter_cpgs_ctrl.xlsx or filter_cpgs_pat.xlsx).
output_file	Character or NULL. Path for the ASM output .xlsx. Auto-named as asm_<sample_type>_<sample_id>.xlsx if NULL.
sample_type	Character. Either "control" or "patient".
verbose	Logical. If TRUE (default), progress messages are written via message(). Set to FALSE or wrap the call in suppressMessages() to silence all output.

Value

A named list with three elements:

asm

Full read-level ASM results as a data.table.

snp_cpg

Per SNP x CpG summary table.

meth_summary

Per CpG allele methylation summary table.

Examples

## Not run: 
  extdata     <- system.file("extdata", package = "ImprintCapASM")
  cpg_snp_tmp <- tempfile(fileext = ".xlsx")

  prepare_cpg_snp_input(
    snp_file     = file.path(extdata, "example_snp.out"),
    meth_file    = file.path(extdata, "example_cgmeth.txt"),
    cpg_ref_file = file.path(extdata, "example_filter_cpgs.xlsx"),
    output_file  = cpg_snp_tmp,
    sample_type  = "control"
  )

  result <- ASM(
    cpg_snp_file     = cpg_snp_tmp,
    sam_file         = file.path(extdata, "example_markdup.bam"),
    filter_cpgs_file = file.path(extdata, "example_filter_cpgs.xlsx"),
    output_file      = tempfile(fileext = ".xlsx"),
    sample_type      = "control"
  )
  head(result$snp_cpg)

## End(Not run)

---

Extract BAM Regions Around SNP Windows

Description

Calls samtools view and samtools sort to subset a whole-genome bisulfite BAM file to the genomic windows defined in a BED file, producing a smaller indexed BAM suitable for input to ASM.

Usage

extract_bam_regions(
  bam_file,
  bed_file,
  output_dir = dirname(bam_file),
  overwrite = FALSE,
  sample_type = c("control", "patient"),
  verbose = TRUE
)

Arguments

bam_file	Character. Path to the input _markdup.bam file (a .bai index must be present alongside it).
bed_file	Character. Path to the BED file produced by prepare_cpg_snp_input.
output_dir	Character. Directory for output files. Defaults to dirname(bam_file).
overwrite	Logical. If FALSE (default), skips extraction when the output BAM already exists.
sample_type	Character. Either "control" or "patient".
verbose	Logical. If TRUE (default), progress messages are written via message(). Set to FALSE or wrap the call in suppressMessages() to silence all output.

Details

Requires samtools to be available on the system PATH. The function calls samtools view -b, samtools sort, and samtools index in sequence. If no .bai index is found alongside bam_file, indexBam is called automatically.

Value

The path to the output BAM file (returned invisibly).

Examples

if (nchar(Sys.which("samtools")) > 0L) {
  extdata <- system.file("extdata", package = "ImprintCapASM")

  out_bam <- extract_bam_regions(
    bam_file    = file.path(extdata, "example_markdup.bam"),
    bed_file    = file.path(extdata, "example_cpg_snp.bed"),
    output_dir  = tempdir(),
    sample_type = "control"
  )
  file.exists(out_bam)
}

---

Prepare CpG-SNP Input Table

Description

Loads a bisulfite SNP .out file and a CpG methylation file, intersects heterozygous SNPs with the 41-DMR CpG panel, and returns a per-sample CpG-SNP pair table ready for input to ASM.

Usage

prepare_cpg_snp_input(
  snp_file,
  meth_file,
  cpg_ref_file,
  output_file = NULL,
  min_depth = 20L,
  window_bp = 60L,
  sample_type = c("control", "patient"),
  verbose = TRUE
)

Arguments

snp_file	Character. Path to a VCF-like file ending in _all.SNPs.out.
meth_file	Character. Path to the gemBS/Bismark CpG methylation table (.cov or .txt).
cpg_ref_file	Character. Path to the CpG panel reference Excel file (filter_cpgs_ctrl.xlsx or filter_cpgs_pat.xlsx).
output_file	Character or NULL. Path for the output .xlsx. Auto-named as cpg_snps_CG_<sample_type>_<sample_id>.xlsx if NULL.
min_depth	Integer. Minimum total SNP read depth. Default 20L.
window_bp	Integer. Window in base pairs around each SNP used to search for CpG positions. Default 60L.
sample_type	Character. Either "control" or "patient".
verbose	Logical. If TRUE (default), progress messages are written via message(). Set to FALSE or wrap the call in suppressMessages() to silence all output.

Value

A data.table of CpG-SNP pairs (returned invisibly). The .xlsx and .bed files are written as side effects.

Examples

extdata    <- system.file("extdata", package = "ImprintCapASM")
  cpg_snp_tmp <- tempfile(fileext = ".xlsx")

  result <- prepare_cpg_snp_input(
    snp_file     = file.path(extdata, "example_snp.out"),
    meth_file    = file.path(extdata, "example_cgmeth.txt"),
    cpg_ref_file = file.path(extdata, "example_filter_cpgs.xlsx"),
    output_file  = cpg_snp_tmp,
    sample_type  = "control"
  )
  head(result)

---

Run the Full ImprintCapASM Pipeline

Description

Convenience wrapper that executes the complete three-step pipeline: (1) prepare_cpg_snp_input, (2) extract_bam_regions, and (3) ASM for every sample in a directory. Controls and patients are always processed independently using their respective filter_cpgs reference files.

Usage

run_pipeline(
  bam_dir,
  snp_dir,
  meth_dir,
  cpg_ref_file,
  output_dir,
  sample_type = c("control", "patient"),
  bed_file,
  min_depth = 20L,
  window_bp = 60L,
  overwrite = FALSE,
  verbose = TRUE
)

Arguments

bam_dir	Character. Path to the directory containing .bam files (and their .bai indices).
snp_dir	Character. Path to the directory containing SNP .out files produced by your bisulfite SNP caller.
meth_dir	Character. Path to the directory containing CpG methylation files (Bismark .cov or similar).
cpg_ref_file	Character. Path to the cohort-matched filter_cpgs reference .xlsx file. Use the file bundled in inst/extdata/filter_cpgs_ctrl.xlsx for controls and inst/extdata/filter_cpgs_pat.xlsx for patients.
output_dir	Character. Directory where all output files are written. Created automatically if it does not exist.
sample_type	Character. Either "control" or "patient". Must match the cohort of samples in bam_dir.
bed_file	Character. Path to the BED file defining the 41 DMR regions used by extract_bam_regions.
min_depth	Integer. Minimum read depth filter passed to prepare_cpg_snp_input. Default 20L.
window_bp	Integer. Window in base pairs around each CpG passed to prepare_cpg_snp_input. Default 60L.
overwrite	Logical. If TRUE, re-runs extraction even when output BAM files already exist. Default FALSE.
verbose	Logical. If TRUE, progress messages are printed to the console via message(). Default TRUE. Set to FALSE or wrap the call in suppressMessages() to silence all output.

Value

Invisibly returns a named list of ASM result objects, one per sample. Each element is the list returned by ASM, containing $asm, $snp_cpg, and $meth_summary.

Examples

## Not run: 
  extdata    <- system.file("extdata", package = "ImprintCapASM")
  cpg_ref    <- file.path(extdata, "example_filter_cpgs.xlsx")
  bed        <- file.path(extdata, "example_cpg_snp.bed")
  output_dir <- tempdir()

  results <- run_pipeline(
    bam_dir      = extdata,
    snp_dir      = extdata,
    meth_dir     = extdata,
    cpg_ref_file = cpg_ref,
    output_dir   = output_dir,
    sample_type  = "control",
    bed_file     = bed
  )
  head(results[[1]]$asm)

## End(Not run)

---