Hello All I am using hisat2 for alignment and RSEM for estimated counts as suggest by a previous post on this site. I need some guidance regarding 1> Does RSEM accepts hisat2 generated transcriptome indexes?
2> rsem-calculate-expression can be used alone without rsem-prepare-reference step and BAM from hisat can be accepted in RSEM count matrix.
I tried for my Q1 here and had error that RSEM was unable to find indexes
reference indexing showed following error. (rsem)> rsem-prepare-reference RNASeq_transcriptome.fa ref rsem-synthesis-reference-transcripts 0 0RNASeq_transcriptome.fa (ASCII code 13), at line 2, position 61!scriptome.fa contains an unknown character, "rsem-synthesis-reference-transcripts ref 0 0 RNASeq_transcriptome.fa" failed! Plase check if you provide correct parameters/options for the pipeline! have used same transcriptome to build hisat2 index successfully
My RNA-seq experiment is 2 Genotypes1control, genotype1treated, Genotypes2control, genotype2treated, >3 replicates-PE each i.e 12 samples. I have reference transcriptome but no GTF/gff files available.
will be very thankful for any suggestion regarding another pipeline or current problem. thank you