bulk_windows_pipeline_setup {FLAMES} | R Documentation

## FLAMES Windows Bulk Pipeline

### Description

An implementation of the FLAMES pipeline designed to run on Windows, or any OS without access to minimap2, for read realignment. This pipeline requires external read alignment, in betwen pipeline calls.

### Usage

    bulk_windows_pipeline_setup(
      annot,
      fastq,
      in_bam = NULL,
      outdir,
      genome_fa,
      downsample_ratio = 1,
      config_file
    )

### Arguments

`annot` | gene annotations file in gff3 format
`fastq` | file path to input fastq file
`in_bam` | optional bam file to use instead of fastq files (skips read alignment step)
`outdir` | directory to store all output files.
`genome_fa` | genome fasta file.
`downsample_ratio` | downsampling ratio if performing downsampling analysis.
`config_file` | JSON configuration file. If specified, `config_file` overrides all configuration parameters

### Details

This function, `bulk_windows_pipeline_setup` is the first step in the 3 step Windows FLAMES bulk pipeline, and should be run first, read alignment undertaken, then `windows_pipline_isoforms` should be run, read realignment performed, and finally `windows_pipeline_quantification` should be run. For each function, besides `bulk_windows_pipeline_setup`, a list `pipeline_variables` is returned, which contains the information required to continue the pipeline. This list should be passed into each function, and updated with the returned list. In the case of `bulk_windows_pipeline_setup`, `pipeline_variables` is the list returned. See the vignette 'Vignette for FLAMES bulk on Windows' for more details.

### Value

a list `pipeline_variables` with the required variables for execution of later Windows pipeline steps. File paths required to perform minimap2 alignment are given in pipeline_variables$return_files. This list should be given as input for `windows_pipeline_isoforms` after minimap2 alignment has taken place; `windows_pipeline_isoforms` is the continuation of this pipeline.

### Examples

    ## example windows pipeline for BULK data. See Vignette for single cell data.

    # download the two fastq files, move them to a folder to be merged together
    temp_path <- tempfile()
    bfc <- BiocFileCache::BiocFileCache(temp_path, ask=FALSE)
    file_url <-
        "https://raw.githubusercontent.com/OliverVoogd/FLAMESData/master/data"
    # download the required fastq files, and move them to new folder
    fastq1 <- bfc[[names(BiocFileCache::bfcadd(bfc, "Fastq1", paste(file_url, "fastq/sample1.fastq.gz", sep="/")))]]
    fastq2 <- bfc[[names(BiocFileCache::bfcadd(bfc, "Fastq2", paste(file_url, "fastq/sample2.fastq.gz", sep="/")))]]
    fastq_dir <- paste(temp_path, "fastq_dir", sep="/") # the downloaded fastq files need to be in a directory to be merged together
    dir.create(fastq_dir)
    file.copy(c(fastq1, fastq2), fastq_dir)
    unlink(c(fastq1, fastq2)) # the original files can be deleted

    # run the FLAMES bulk pipeline setup
    #pipeline_variables <- bulk_windows_pipeline_setup(annot=system.file("extdata/SIRV_anno.gtf", package="FLAMES"),
    #                   fastq=fastq_dir,
    #                   outdir=tempdir(), genome_fa=system.file("extdata/SIRV_genomefa.fasta", package="FLAMES"),
    #                   config_file=system.file("extdata/SIRV_config_default.json", package="FLAMES"))
    # read alignment is handled externally (below downloads aligned bam for example)
    # genome_bam <- paste0(temp_path, "/align2genome.bam")
    # file.rename(bfc[[names(BiocFileCache::bfcadd(bfc, "Genome BAM", paste(file_url, "align2genome.bam", sep="/")))]], genome_bam)
    #
    # genome_index <- paste0(temp_path, "/align2genome.bam.bai")
    # file.rename(bfc[[names(BiocFileCache::bfcadd(bfc, "Genome BAM Index", paste(file_url, "align2genome.bam.bai", sep="/")))]], genome_index)
    # pipeline_variables$genome_bam = genome_bam
    #
    # # run the FLAMES bulk pipeline find isoforms step
    # pipeline_variables <- windows_pipeline_isoforms(pipeline_variables)
    #
    # # read realignment is handled externally
    # realign_bam <- paste0(temp_path, "/realign2genome.bam")
    # file.rename(bfc[[names(BiocFileCache::bfcadd(bfc, "Realign BAM", paste(file_url, "realign2transcript.bam", sep="/")))]], realign_bam)
    #
    # realign_index <- paste0(temp_path, "/realign2genome.bam.bai")
    # file.rename(bfc[[names(BiocFileCache::bfcadd(bfc, "Realign BAM Index", paste(file_url, "realign2transcript.bam.bai", sep="/")))]], realign_index)
    # pipeline_variables$realign_bam <- realign_bam
    #
    # # finally, quantification, which returns a Summarized Experiment object
    # se <- windows_pipeline_quantification(pipeline_variables)

[Package FLAMES version 1.0.2 Index]