Pipeline for Single Cell Data (deprecated)

This function is deprecated. Please use [SingleCellPipeline()] instead.

Usage

sc_long_pipeline(
  annotation,
  fastq,
  outdir,
  genome_fa,
  minimap2 = NULL,
  barcodes_file = NULL,
  expect_cell_number = NULL,
  config_file = NULL
)

Arguments

annotation

The file path to the annotation file in GFF3 format

fastq

The file path to input fastq file

outdir

The path to directory to store all output files.

genome_fa

The file path to genome fasta file.

minimap2

Path to minimap2, optional.

barcodes_file

The file with expected cell barcodes, with each barcode on a new line.

expect_cell_number

The expected number of cells in the sample. This is used if barcodes_file is not provided. See BLAZE for more details.

config_file

File path to the JSON configuration file.

Value

A SingleCellPipeline object containing the transcript counts.

See also

SingleCellPipeline for the new pipeline interface, BulkPipeline for bulk long data, MultiSampleSCPipeline for multi sample single cell pipelines.

Examples

outdir <- tempfile()
dir.create(outdir)
bc_allow <- file.path(outdir, "bc_allow.tsv")
genome_fa <- file.path(outdir, "rps24.fa")
R.utils::gunzip(
  filename = system.file("extdata", "bc_allow.tsv.gz", package = "FLAMES"),
  destname = bc_allow, remove = FALSE
)
R.utils::gunzip(
  filename = system.file("extdata", "rps24.fa.gz", package = "FLAMES"),
  destname = genome_fa, remove = FALSE
)
sce <- FLAMES::sc_long_pipeline(
  genome_fa = genome_fa,
  fastq = system.file("extdata", "fastq", "musc_rps24.fastq.gz", package = "FLAMES"),
  annotation = system.file("extdata", "rps24.gtf.gz", package = "FLAMES"),
  outdir = outdir,
  barcodes_file = bc_allow,
  config_file = FLAMES::create_config(
    outdir,
    pipeline_parameters.demultiplexer = "flexiplex"
  )
)
#> ℹ Writing configuration to: /tmp/Rtmp4nGYdi/filebc442770b76d/config_file_48196.json
#> Configured steps: 
#> 	barcode_demultiplex: TRUE
#> 	genome_alignment: TRUE
#> 	gene_quantification: TRUE
#> 	isoform_identification: TRUE
#> 	read_realignment: TRUE
#> 	transcript_quantification: TRUE
#> samtools not found, will use Rsamtools package instead
#> ── Running step: barcode_demultiplex @ Fri Jun 26 08:25:45 2026 ────────────────
#> Using flexiplex for barcode demultiplexing.
#> Loading known barcodes from /tmp/Rtmp4nGYdi/filebc442770b76d/bc_allow.tsv
#> Number of known barcodes: 143
#> FLEXIPLEX 1.02.6
#> Setting max flanking sequence edit distance to 8
#> Setting number of threads to 8
#> Search pattern:
#> primer: CTACACGACGCTCTTCCGATCT
#> CB: NNNNNNNNNNNNNNNN
#> UB: NNNNNNNNNNNN
#> polyT: TTTTTTTTT
#> CB:Z: tag field: CB
#> Processing file: /__w/_temp/Library/FLAMES/extdata/fastq/musc_rps24.fastq.gz
#> Searching for barcodes...
#> Number of reads processed: 393
#> Number of reads where at least one barcode was found: 368
#> Number of chimera reads: 1
#> All done!
#> Reads	Barcodes
#> 10	2
#> 9	2
#> 8	5
#> 7	4
#> 6	3
#> 5	7
#> 4	14
#> 3	14
#> 2	29
#> 1	57
#> ── Running step: genome_alignment @ Fri Jun 26 08:25:45 2026 ───────────────────
#> Creating junction bed file from GFF3 annotation.
#> Aligning sample /tmp/Rtmp4nGYdi/filebc442770b76d/matched_reads.fastq.gz -> /tmp/Rtmp4nGYdi/filebc442770b76d/align2genome.bam
#> Warning: samtools not found, using Rsamtools instead, this could be slower and might fail for large BAM files.
#> Sorting BAM files by genome coordinates with 8 threads...
#> Indexing bam files
#> ── Running step: gene_quantification @ Fri Jun 26 08:25:46 2026 ────────────────
#> 08:25:46 AM Fri Jun 26 2026 quantify genes 
#> Using BAM(s): /tmp/Rtmp4nGYdi/filebc442770b76d/align2genome.bam
#> ── Running step: isoform_identification @ Fri Jun 26 08:25:46 2026 ─────────────
#> ── Running step: read_realignment @ Fri Jun 26 08:25:46 2026 ───────────────────
#> Checking for fastq file(s) /__w/_temp/Library/FLAMES/extdata/fastq/musc_rps24.fastq.gz
#> 	files found
#> Checking for fastq file(s) /tmp/Rtmp4nGYdi/filebc442770b76d/matched_reads.fastq.gz
#> 	files found
#> Checking for fastq file(s) /tmp/Rtmp4nGYdi/filebc442770b76d/matched_reads_dedup.fastq.gz
#> 	files found
#> Realigning sample /tmp/Rtmp4nGYdi/filebc442770b76d/matched_reads_dedup.fastq.gz -> /tmp/Rtmp4nGYdi/filebc442770b76d/realign2transcript.bam
#> Warning: samtools not found, using Rsamtools instead, this could be slower and might fail for large BAM files.
#> Sorting BAM files by 8 with CB threads...
#> ── Running step: transcript_quantification @ Fri Jun 26 08:25:47 2026 ──────────
#> Pipeline saved to /tmp/Rtmp4nGYdi/filebc442770b76d/pipeline.rds