coverage_norm {dasper} | R Documentation

## Processing coverage

### Description

The set of functions prefixed with "coverage_" are used to process coverage data. They are designed to be run after you have processed your junctions in the order `coverage_norm`, `coverage_score`. Or, alternatively the wrapper function `coverage_process` can be used to run the 2 functions stated above in one go. For more details of the individual functions, see "Details".

### Usage

    coverage_norm(
      junctions,
      ref,
      unannot_width = 20,
      coverage_paths_case,
      coverage_paths_control,
      coverage_chr_control = NULL,
      load_func = .coverage_load,
      bp_param = BiocParallel::SerialParam(),
      norm_const = 1
    )

    coverage_process(
      junctions,
      ref,
      unannot_width = 20,
      coverage_paths_case,
      coverage_paths_control,
      coverage_chr_control = NULL,
      load_func = .coverage_load,
      bp_param = BiocParallel::SerialParam(),
      norm_const = 1,
      score_func = .zscore,
      ...
    )

    coverage_score(junctions, coverage, score_func = .zscore, ...)

### Arguments

`junctions` | junction data as a RangedSummarizedExperiment-class object.
`ref` | either path to gtf/gff3 or object of class TxDb-class.
`unannot_width` | integer scalar determining the width of the region to obtain coverage from when the end of of a junction does not overlap an existing exon.
`coverage_paths_case` | paths to the BigWig files containing the coverage of your case samples. Must be the same length and order to the samples in `junctions`.
`coverage_paths_control` | paths to the BigWig files
`coverage_chr_control` | either "chr" or "no_chr", indicating the chromosome format of control coverage data. Only required if the chromosome format of the control BigWig files is different to that of your junctions.
`load_func` | a function to use to load coverage. Currently only for internal use to increase testing speed.
`bp_param` | a BiocParallelParam-class instance denoting whether to parallelise the loading of coverage across BigWig files.
`norm_const` | numeric scaler to add to the normalisation coverage to avoid dividing by 0s and resulting NaN or Inf values.
`score_func` | function to score junctions by their abnormality. By default, will use a z-score but can be switched to a user-defined function. This function must take as input an `x` and `y` argument, containing case and control counts respectively. This must return a numeric vector equal to the length of `x` with elements corresponding to a abnormality of each junction.
`...` | additional arguments passed to `score_func`.
`coverage` | list containing normalised coverage data that is outputted from coverage_norm.

### Details

`coverage_process` wraps all "coverage_" prefixed functions in dasper. This is designed to simplify processing of the coverage data for those familiar or uninterested with the intermediates.

`coverage_norm` obtains regions of interest for each junction where coverage disruptions would be expected. These consist of the intron itself the overlapping exon definitions (if ends of junctions are annotated), picking the shortest exon when multiple overlap one end. If ends are unannotated, `coverage_norm` will use a user-defined width set by `unannot_width`. Then, coverage will be loaded using megadepth and normalised to a set region per junction. By default, the boundaries of each gene associated to a junction are used as the region to normalise to.

`coverage_score` will score disruptions in the coverage across the intronic/exonic regions associated with each junction. This abnormality score generated by `score_func` operates by calculating the deviation of the coverage in patients to a coverage across the same regions in controls. Then, for each junction it obtains the score of the region with the greatest disruption.

### Value

junctions as SummarizedExperiment object with additional `assays` named "coverage_region" and "coverage_score". "coverage_region" labels the region of greatest disruption (1 = exon_start, 2 = exon_end, 3 = intron) and "coverage_score" contains the abnormality scores of the region with the greatest disruption.

### Functions

- `coverage_norm`: Load and normalise coverage from RNA-sequencing data

- `coverage_score`: Score coverage by their abnormality

### Examples

    ##### Set up txdb #####

    # use GenomicState to load txdb (GENCODE v31)
    ref <- GenomicState::GenomicStateHub(
        version = "31",
        genome = "hg38",
        filetype = "TxDb"
    )[[1]]

    ##### Set up BigWig #####

    # obtain path to example bw on recount2
    bw_path <- recount::download_study(
        project = "SRP012682",
        type = "samples",
        download = FALSE
    )[[1]]

    ##### junction_process #####

    junctions_processed <- junction_process(
        junctions_example,
        ref,
        types = c("ambig_gene", "unannotated"),
    )

    ##### coverage_norm #####

    coverage_normed <- coverage_norm(
        junctions_processed,
        ref,
        unannot_width = 20,
        coverage_paths_case = rep(bw_path, 2),
        coverage_paths_control = rep(bw_path, 2)
    )

    ##### coverage_score #####

    junctions <- coverage_score(junctions_processed, coverage_normed)

    ##### coverage_process #####

    # this wrapper will obtain coverage scores identical to those
    # obtained through running the individual wrapped functions shown below
    junctions_w_coverage <- coverage_process(
        junctions_processed,
        ref,
        coverage_paths_case = rep(bw_path, 2),
        coverage_paths_control = rep(bw_path, 3)
    )

    # the two objects are equivalent
    all.equal(junctions_w_coverage, junctions, check.attributes = FALSE)

[Package dasper version 1.2.0 Index]