Abnormal spliced/unspliced percentages using RNA velocity tutorial

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Angela Ling

Aug 3, 2022, 3:29:00 PM8/3/22
to kallisto and applications

Thank you for creating such an innovative package. I am trying to follow the kb-python RNA velocity tutorial using my 10x cell-hashed data. I was able to download the pre-built mouse index files and create a loom file for my data. 

This is what I ran:

 kb ref -i index.idx -g t2g.txt -f1 cdna.fa -f2 intron.fa -c1 cdna_t2c.txt -c2 intron_t2c.txt --lamanno Mus_musculus.GRCm38.dna.primary_assembly.fa.gz Mus_musculus.GRCm38.98.gtf.gz 

 kb count --loom -i index.idx -g t2g.txt -x 10xv3 -o D3repI_fastq -c1 cDNA_tx_to_capture.txt -c2 introns_tx_to_capture.txt --workflow lamanno --filter bustools -t 6 sample1_L001_R1_001.fastq.gz  sample1_L001_R2_001.fastq.gz sample1_L002_R1_001.fastq.gz sample1_L002_R2_001.fastq.gz sample1_L003_R1_001.fastq.gz sample1_L003_R2_001.fastq.gz sample1_L004_R1_001.fastq.gz sample1__L004_R2_001.fastq.gz

When I check the spliced proportions in scvelo, I get 41% spliced and 41% unspliced.
Here are the values I get when I check the fraction of unspliced and spliced counts: 
[nan 0.5 0.5 ... 0.5 0.5 0.5] 

[nan 0.5 0.5 ... 0.5 0.5 0.5]

I was wondering why I might be getting these strange results. 

Thanks so much for the help!

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