Hi,

I am using linkedTxome function in tximeta in order to use a local transcriptome database. However, in some random runs, the GTF file is being downloaded from a FTP server which causes errors when the device is not connected to the internet.

Here is the code I use:

  s <- data.frame(files = quant, names = sample, label = label)

  # setting BiocFileCache directory for accessing and saving TxDb sqlite files
  BFCdir <- file.path(dirname(spath), "BFC")
  setTximetaBFC(BFCdir)

  makeLinkedTxome(indexDir = index_dir,
          source = "gencode",
          organism = "human",
          release = "v31",
          genome = "GRCh38",
          fasta = fasta_file,
          gtf = gtf_path,
          write = FALSE)

  txi.methname <- tximeta(s, type = ftype, useHub = FALSE)

And the error message:

NOTE: linkedTxome with source='GENCODE', set useHub=FALSE in tximeta
to avoid download of reference txome from AnnotationHub.
alternatively use a different string for source argument
linkedTxome was different than one in bfc, replacing
importing quantifications
reading in files with read_tsv
1 2 3 4 5 6 7 8 9 10
found matching transcriptome:
[ GENCODE - Homo sapiens - release 31 ]
building TxDb with 'GenomicFeatures' package
Import genomic features from the file as a GRanges object ... trying URL 'ftp://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_31/gencode.v31.annotation.gtf.gz'
Error in download.file(resource(con), destfile) :
  cannot open URL 'ftp://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_31/gencode.v31.annotation.gtf.gz'
Calls: getTxiResults2 ... import -> import -> import -> .local -> download.file
In addition: Warning message:
In download.file(resource(con), destfile) :
  cannot open URL 'ftp://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_31/gencode.v31.annotation.gtf.gz': FTP status was '421 Service not available, closing control connection'
Execution halted

I was wondering if there is a way to ensure the local GTF file is going to be used instead of attempting to download it from a FTP server.

Thanks in advance.

The very first place you should check, before coming here to ask, is the help page for the function you are using. Which says

Arguments:

indexDir: the local path to the Salmon index

  source: the source of transcriptome (e.g. "de-novo"). Note: if you
          specify "GENCODE" or "Ensembl", this will trigger behavior by
          tximeta that may not be desired: e.g. attempts to download
          canonical transcriptome data from AnnotationHub (unless
          useHub=FALSE when running tximeta) and parsing of Ensembl GTF
          using ensembldb (which may fail if the GTF file has been
          modified). For transcriptomes that are defined by local GTF
          files, it is recommended to use the terms "LocalGENCODE" or
          "LocalEnsembl"

Which I believe clearly answers your question.