Create SingleCellExperiment
object from FLAMES
output folder
Source: R/sc_long_pipeline.R
create_sce_from_dir.Rd
Create SingleCellExperiment
object from FLAMES
output folder
Arguments
- outdir
The folder containing
FLAMES
output files- annotation
the annotation file that was used to produce the output files
- quantification
(Optional) the quantification method used to generate the output files (either "FLAMES" or "Oarfish".). If not specified, the function will attempt to determine the quantification method.
Value
a list of SingleCellExperiment
objects if multiple transcript matrices were
found in the output folder, or a SingleCellExperiment
object if only one were found
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
)
annotation <- system.file("extdata", "rps24.gtf.gz", package = "FLAMES")
sce <- sc_long_pipeline(
genome_fa = genome_fa,
fastq = system.file("extdata", "fastq", "musc_rps24.fastq.gz", package = "FLAMES"),
annotation = annotation,
outdir = outdir,
barcodes_file = bc_allow,
config_file = create_config(outdir, oarfish_quantification = FALSE)
)
#> Writing configuration parameters to: /tmp/RtmpgpEV0i/file25bfae1bd27/config_file_9663.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
#> FLEXIPLEX 0.96.2
#> Setting max barcode edit distance to 2
#> Setting max flanking sequence edit distance to 8
#> Setting read IDs to be replaced
#> Setting number of threads to 8
#> Search pattern:
#> primer: CTACACGACGCTCTTCCGATCT
#> BC: NNNNNNNNNNNNNNNN
#> UMI: NNNNNNNNNNNN
#> polyT: TTTTTTTTT
#> Setting known barcodes from /tmp/RtmpgpEV0i/file25bfae1bd27/bc_allow.tsv
#> Number of known barcodes: 143
#> 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 reads with exactly one barcode match: 364
#> Number of chimera reads: 1
#> All done!
#> Running step: genome_alignment
#> Creating junction bed file from GFF3 annotation.
#> Aligning sample /tmp/RtmpgpEV0i/file25bfae1bd27/matched_reads.fastq -> /tmp/RtmpgpEV0i/file25bfae1bd27/align2genome.bam
#> Your fastq file appears to have tags, but you did not provide the -y option to minimap2 to include the tags in the output.
#> 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
#> 03:22:27 AM Wed May 21 2025 quantify genes
#> Found genome alignment file(s): align2genome.bam
#> Running step: isoform_identification
#> Import genomic features from the file as a GRanges object ...
#> OK
#> Prepare the 'metadata' data frame ...
#> OK
#> Make the TxDb object ...
#> Warning: genome version information is not available for this TxDb object
#> OK
#> Running step: read_realignment
#> Checking for fastq file(s) /__w/_temp/Library/FLAMES/extdata/fastq/musc_rps24.fastq.gz
#> files found
#> Checking for fastq file(s) /tmp/RtmpgpEV0i/file25bfae1bd27/matched_reads.fastq
#> files found
#> Checking for fastq file(s) /tmp/RtmpgpEV0i/file25bfae1bd27/matched_reads_dedup.fastq
#> files found
#> Realigning sample /tmp/RtmpgpEV0i/file25bfae1bd27/matched_reads_dedup.fastq -> /tmp/RtmpgpEV0i/file25bfae1bd27/realign2transcript.bam
#> Your fastq file appears to have tags, but you did not provide the -y option to minimap2 to include the tags in the output.
#> 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: transcript_quantification
#> 03:22:28 AM Wed May 21 2025 quantify transcripts
#> Warning: Annotation in GFF format may cause errors. Please consider using GTF formats.
#> Found realignment file(s): realign2transcript.bam
#> Pipeline saved to /tmp/RtmpgpEV0i/file25bfae1bd27/pipeline.rds
sce_2 <- create_sce_from_dir(outdir, annotation)