I am currently trying to use kallisto and IGV to visualize reads from a single-cell RNASeq experiment. I'm interested in non-coding RNA, but I would still like to see how the reads align to each chromosome. Currently, I can visualize how the reads align to each individual non-coding transcript by aligning to the ensembl non-coding RNA fasta. But when I tried implementing --genomebam and the GTF file from ensembl, I stopped seeing my reads when I aligned to the genome fasta. My code below successfully generates my transcript index:
! kallisto index -i transcripts.idx Homo_sapiens.GRCh38.ncrna.fa.gz
But then I try the below code to generate a bam file to use in IGV, such that I see how the reads align to the genome...
! kallisto quant --genomebam --gtf Homo_sapiens.GRCh38.99.gtf.gz -i transcripts.idx -o output 1.fastq 2.fastq
I get a bam file successfully, but I don't see any alignments in IGV.
For what it's worth, I get the below warning:
Warning: 173489 transcripts were defined in GTF file, but not in the index
I've read in the kallisto manual that I may also need a chromosome number/size file, but I don't know how to get such a file.
Thanks so much