Hi I am runing into very frustrating issue. I have RNA seq pair end samples that i am processing with Rsubread package. For the sample nr1 test everyhting worked fine and i obtained feature counts which look reasonable. However, after alignment of sample nr 2, attempt to perform feature counting returns this message
nput files : 1 BAM file || || || || OV2_Sample_aligned.bam || || || || Paired-end : yes || || Count read pairs : yes || || Annotation : gencode.v38.annotation.gtf (SAF) || || Dir for temp files : . || || Threads : 1 || || Level : meta-feature level || || Multimapping reads : counted || || Multi-overlapping reads : not counted || || Min overlapping bases : 1 || || || \============================================================================//
i dont have a slightest idea why the program says the file is SAF ( it is GTF from encode and it worked for the first sample)
i used relatively straight forward code
''library(Rsubread)
bam_file <- "path/aligned_reads.bam" gtf_file <- "path/to/annotation.gtf"
counts <- featureCounts(files = bam_file, annot.ext = gtf_file, isPairedEnd = TRUE)"
what could be wrong???? GTF file is absolutely fine i did check it in text editor.
Any suggestions?