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Read the trace files embedded in a Sequencher project

read_spf.Rd

Extracts every chromatogram stored in a Sequencher project file (.SPF), with the base calls, quality scores and peak positions that go with it.

Usage

read_spf(path, deleted = FALSE)

Arguments

path

Path to a Sequencher project file.

deleted

If TRUE, also return reads deleted from the project whose data is still in the file.

Value

A list with one element per read, named after the reads. Where names repeat, ignoring case, later copies get a suffix (_2, _3, ...); reads that are not deleted are numbered first, so their names are the same whatever deleted is. These are the names spf_to_abif() gives its files and read_spf_contigs() gives contig members. Each element is a list with

  • name: the file name the read was imported under, as shown in the project. Names can repeat: a file imported twice gives two reads with one name. A deleted read whose name has been overwritten is named unnamed_read_<n>, where n is its position among all reads in the file;

  • bases: base calls as IUPAC codes, with - for a gap;

  • quality: Phred quality scores, one per base, or an empty vector if the project stores none for the read;

  • peaks: the scan position of each base call, counted from 0 as in the ABIF PLOC tags, so base i sits at row peaks[i] + 1 of traces;

  • traces: an integer matrix of processed trace intensities with columns A, C, G and T, one row per scan;

  • deleted: TRUE for a read deleted from the project, FALSE otherwise, and NA if the file has no object table to tell them apart.

Details

The traces are the processed (analysed) channels, the data an .ab1 file holds in its DATA9–DATA12 tags. The sequencer's software has already subtracted the baseline, separated the dye signals and corrected for mobility, so these are the curves a trace viewer shows. The raw detector signal (DATA1–DATA4) is not stored in the project.

The bases, quality scores and peak positions are the ones stored in the project, so they include any edits made in Sequencher. The project keeps no copy of the original calls and no flag marking which reads were edited, so edited and unedited reads cannot be told apart. In a project where 150 reads could be checked against their original files, two had been edited, each by deleting one base.

Sequencher can leave the data of deleted reads in the file. A read counts as deleted when no object table in the file lists it; spf_format describes the tables. If the file has none, every read is returned, with a warning.

The .SPF format is undocumented. spf_format describes how it was decoded and checked, and which parts are not read yet.

Examples

spf <- "project.SPF"
if (file.exists(spf)) {
  reads <- read_spf(spf)
  names(reads)
  matplot(reads[[1]]$traces[1:500, ], type = "l", lty = 1)
}