Nothing new in this release; just some internal checks and maintenance.

• Add 10xv4
• --exact-barcodes in kb count to "correct" barcodes to an on-list using only exact matches (i.e. no mismatches permitted)
• bustools binary updated to 0.45.0
• Allow kb count -g None (i.e. not supplying a t2g.txt file) in which case a synthetic one is generated with each target/transcript being its own gene.

Slightly modified the --h5ad to lower RAM spike when dealing with a large number of genes

Updates since version 0.28.2:

Major:

• Upgraded kallisto to 0.51.1 and bustools to 0.44.1
• Added lr-kallisto (--long) option, and enabling k>31
• Added kb extract
• Added various kallisto binaries (w/ and w/o optimizations; w/ and w/o long k-mer sizes)

Other:

• Allow -i NONE in kb ref to create t2g+fasta but no index
• Various bug fixes (pandas version dependency, adata.X in nac containing total matrix, summing matrices not mishandling scientific notation, etc.)
• Ended support for python 3.7

Fix: Fixed how nascent/mature/ambiguous matrices are stored in anndata/loom files

Fix: The old lamanno workflow (for legacy purposes) should work now.

Anndata/loom files now have nascent/mature layers rather than unspliced/spliced layers.

--workflow=custom can take in multiple FASTA inputs

Allow --d-list to have comma-separated multiple FASTA files with URLs

Command-line options menu cleaned up a bit

Implements all the updates detailed in protocols paper: https://doi.org/10.1101/2023.11.21.568164

• kallisto version 0.50.1
• bustools version 0.43.1

General

• Bumped ngs-tools>=1.7.3.

ref

• [DEPRECATION] Split index generation using -n has been fully deprecated. (Thanks to @amcdavid for catching a bug)

count

• Fixed a minor issue with --workflow kite:10xFB, where bustools project would be called before bustools correct (the order should be opposite). This fix required a bump to the ngs-tools dependency.
• Support for --workflow lamanno for -x smartseq3.
• [DEPRECATION] Counting using split indices by providing a comma-delimited list to -i has been fully deprecated.
• Support for whitelist (-w option) for bulk, smartseq2 and smartseq3 technologies.
• Added support for -x 10XV3_ULTIMA.

count

• Whitelist for technology -x BDWTA is now provided.

General

• [DEPRECATION] Support for split indices (with the -n option) will be deprecated in the next major release. It is now recommended to use --include-attribute and --exclude-attribute options, similar to Cellranger's mkref options (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/advanced/references), to kb ref to reduce index size and memory usage.

ref

• A remote URL may be provided as the fasta (genomic FASTA) and/or gtf (gene annotation GTF) arguments. Support from ngs_tools 1.5.13.
• GTF is now allowed to have 0-length segments (pachterlab/kallisto#340).

count

• [DEPRECATION] Technology SMARTSEQ is now deprecated. All future uses should use BULK, SMARTSEQ2 or SMARTSEQ3.
• Genes that do not have a gene name will now have their gene IDs in the gene_name column (or the adata.var_names if --gene-names is used).
• Support for --workflow lamanno for -x BULK and -x SMARTSEQ2 technologies.