Oarfish quantifies long reads (Oxford Nanopore and PacBio) against a
transcriptome. For every sample it writes, next to a common output prefix,
<prefix>.quant (transcript name, length and estimated read count),
<prefix>.ambig_info.tsv (reads that map to the transcript alone and reads
shared with others), <prefix>.meta_info.json and, with
--num-bootstraps, <prefix>.infreps.pq (the bootstrap replicates as a
Parquet table). read_oarfish() reads them like read_salmon() reads
Salmon output: an IsoformicExperiment with counts, length and
abundance assays, and, with inferential = TRUE, the uncertainty tables
of add_uncertainty().
Usage
read_oarfish(
paths,
annot_path,
col_data = NULL,
experiment_name = NULL,
ignore_after_bar = TRUE,
drop_unannotated = FALSE,
inferential = FALSE
)Arguments
- paths
Output prefixes of Oarfish (the value of
-o), paths to the.quantfiles, or directories holding a single.quantfile. Names, when present, are the sample IDs; otherwise the prefix is used.- annot_path
Annotation directory or GFF3/GTF file (see
prepare_isoformic_annotation()).- col_data
Optional data frame of sample metadata whose first column (or a
sample_idcolumn) matches the sample IDs. WhenNULL, a minimal table with justsample_idis created.- experiment_name
Optional experiment name.
- ignore_after_bar
Logical. Keep only the part of transcript names before the first
|(GENCODE FASTA headers used as Salmon references).- drop_unannotated
Logical. Drop transcripts absent from the annotation (with a message) instead of failing.
- inferential
Logical. Read the bootstrap replicates and the ambiguity counts and compute the uncertainty tables with
add_uncertainty(). The samples must have been quantified with--num-bootstraps.
Details
Oarfish does not write TPM; the abundance assay is computed from the
read counts and the transcript lengths (reads per base, scaled to one
million). The ambiguity counts of .ambig_info.tsv play the role of the
equivalence classes of Salmon in equivalence_support(): unique_fraction
is the share of reads mapped to the transcript alone, while n_classes is
not available. The functions read_inferential_replicates(),
equivalence_support(), salmon_run_qc() and add_uncertainty() accept
Oarfish prefixes as paths, but not together with Salmon directories.
Examples
prefixes <- file.path(
system.file("extdata", "oarfish", package = "isoformic"),
c("control_1", "control_2", "treated_1")
)
names(prefixes) <- basename(prefixes)
annot <- system.file(
"extdata", "example_experiment", "annotation",
package = "isoformic"
)
iso <- read_oarfish(prefixes, annot_path = annot, inferential = TRUE)
iso
#> <IsoformicExperiment> isoformic_382cc0e633
#> annotation : gencode_v33_example_subset (69 genes, 892 transcripts, 7,032 exons)
#> samples : 3 [oarfish_num_processed, oarfish_num_mapped, oarfish_percent_mapped, oarfish_replicate_type, oarfish_num_replicates, oarfish_optimiser, oarfish_alignment_source, oarfish_prob_model, oarfish_qc_flag]
#> transcripts: 72 assayed from 3 genes
#> assays : abundance, counts, length
#> gene assays: derived from transcript assays on demand
#> DE results : <none>
head(tx_uncertainty(iso))
#> # A tibble: 6 × 18
#> transcript_id gene_id gene_name transcript_name transcript_type mean_count
#> <chr> <chr> <chr> <chr> <chr> <dbl>
#> 1 ENST00000512351.5 ENSG00… RBPJ RBPJ-222 protein_coding 59.9
#> 2 ENST00000510778.6 ENSG00… RBPJ RBPJ-218 protein_coding 13.7
#> 3 ENST00000506956.5 ENSG00… RBPJ RBPJ-213 protein_coding 21.4
#> 4 ENST00000512671.5 ENSG00… RBPJ RBPJ-223 protein_coding 1.19
#> 5 ENST00000345843.7 ENSG00… RBPJ RBPJ-203 protein_coding 8.92
#> 6 ENST00000342295.5 ENSG00… RBPJ RBPJ-201 protein_coding 41.3
#> # ℹ 12 more variables: assessed <lgl>, mean_inf_rv <dbl>, max_inf_rv <dbl>,
#> # mean_inf_cv <dbl>, uncertain <lgl>, min_sibling_cor <dbl>,
#> # n_ambiguous_partners <int>, ambiguity_group <chr>, n_classes <dbl>,
#> # n_fragments <dbl>, n_unique_fragments <dbl>, unique_fraction <dbl>