I have a total RNAseq dataset that I aligned using STAR producing BAM files (sorted by coordinates). I am now trying to get counts for the lncRNA sequences using htseq-count, with the command:
htseq-count -m intersection-nonempty -f bam -i gene_id -r pos -s reverse ./OVCA429_Aligned.sortedByCoord.out.bam ./GRCh38_latest_genomic.gtf > ./OVCA429.counts.txt
This is the output:
[E::idx_find_and_load] Could not retrieve index file for './OVCA429_587680_1_1.rnaseq.fastqAligned.sortedByCoord.out.bam' 100000 GFF lines processed. 200000 GFF lines processed. 300000 GFF lines processed. 400000 GFF lines processed. ...(truncated output here)... 3775001 GFF lines processed. [E::idx_find_and_load] Could not retrieve index file for './OVCA429_587680_1_1.rnaseq.fastqAligned.sortedByCoord.out.bam' 100000 alignment records processed. 200000 alignment records processed. 300000 alignment records processed. ...(truncated output here)... Output text file: __no_feature 20368875 __ambiguous 0 __too_low_aQual 0 __not_aligned 0 __alignment_not_unique 2634439
I've read through the htseq-count literature but it never asks to include an index file, therefore i'm not sure how to deal with this problem. Any help/ suggestions would be welcome! Thank you, in advance.