Lamanno workflow with SMARTSEQ2 data

[reliscu]

Info:

> uname -a
Linux gpu5 5.15.0-88-generic #98-Ubuntu SMP Mon Oct 2 15:18:56 UTC 2023 x86_64 x86_64 x86_64 GNU/Linux

> kallisto version
kallisto, version 0.50.1

My command:

kb count \
  -i "${index_dir}/index_lamanno.idx" \
  -g "${index_dir}/t2g_lamanno.txt" \
  -x SMARTSEQ2 \
  -o "${outdir}/processed/kallisto/VIS" \
  -c1 "${index_dir}/cdna_t2c_lamanno.txt" \
  -c2 "${index_dir}/intron_t2c_lamanno.txt" \
  --workflow lamanno \
  -w NONE \
  --parity paired \
  --h5ad \
  -t 20 -m 256G \
  --batch "${outdir}/batch_test.txt"

Outputs:

[2024-12-08 11:54:25,192]    INFO [count_lamanno] Using index /mnt/lareaulab/reliscu/data/GENCODE/GRCm39/kallisto_index/index_lamanno.
idx to generate BUS file to /mnt/lareaulab/reliscu/projects/protein_prediction/data/BICCN/mouse_CTX_HPF/processed/kallisto/VIS from
[2024-12-08 11:54:25,192]    INFO [count_lamanno]         /mnt/lareaulab/reliscu/projects/protein_prediction/data/BICCN/mouse_CTX_HPF/processed/kallisto/VIS/tmp/tmp3cb093xy
[2024-12-08 11:56:02,573]    INFO [count_lamanno] Sorting BUS file /mnt/lareaulab/reliscu/projects/protein_prediction/data/BICCN/mouse_CTX_HPF/processed/kallisto/VIS/output.bus to /mnt/lareaulab/reliscu/projects/protein_prediction/data/BICCN/mouse_CTX_HPF/processed/kallisto/VIS/tmp/output.s.bus

[2024-12-08 11:57:33,054]    INFO [count_lamanno] Inspecting BUS file /mnt/lareaulab/reliscu/projects/protein_prediction/data/BICCN/mouse_CTX_HPF/processed/kallisto/VIS/tmp/output.s.bus
[2024-12-08 11:57:34,169]   ERROR [count_lamanno] Error: File not found NONE
Usage: bustools inspect [options] sorted-bus-file

I'm not sure how to proceed in order to overcome this issue. Any help would be greatly appreciated.

[Yenaled]

First, remove -w NONE

Second, lamanno is the old workflow; to use it, you might want to try installing kb-python version 0.27.3 or earlier if the current version doesn’t work.

[reliscu]

Hi @Yenaled. Thanks for getting back to me. If I try it without the -w NONE flag, I get an error that using the SMARTSEQ2 workflow requires that flag.

But per your suggestion, I will try an earlier version and see if that works.

[Yenaled]

You could just use -x BULK