Quick start

• Spl-IsoQuant is installed from GitHub:

  git clone https://github.com/algbio/spl-IsoQuant.git 
  cd spl-IsoQuant
  git checkout latest
  pip install -e .
  

Installation typically takes no more than a few minutes.

• Spl-IsoQuant is not distributed via PyPI or bioconda and will not be published there. The original IsoQuant provides the same functionality and is available from both:

  pip install isoquant
  
  conda create -c conda-forge -c bioconda -n isoquant python=3.12 isoquant
  

These install the isoquant executable, not splisoquant. If you install this way, use isoquant in place of splisoquant in all commands below.

• If running simply from the source archive you will need:
• Python3 (3.8 or higher)
• gffutils
• pysam
• biopython
• pyfaidx
• ssw-py
• editdistance
• numba
• mappy
• intervaltree
• xgboost
• scikit-learn

and some other common Python libraries to be installed. See requirements.txt for details.

You will also need to have minimap2 and samtools to be in your $PATH variable.

• All required Python libraries can be installed via:

  pip install -r requirements.txt

• Verify your installation by running (typically takes less than 1 minute):

  splisoquant --test
  

• To run Spl-IsoQuant on raw FASTQ/FASTA files, use the following command

  splisoquant --reference /PATH/TO/reference_genome.fasta \
  --genedb /PATH/TO/gene_annotation.gtf \
  --fastq /PATH/TO/sample1.fastq.gz /PATH/TO/sample2.fastq.gz \
  --data_type (assembly|pacbio_ccs|nanopore) -o OUTPUT_FOLDER
  

  For example, using the toy data provided within this repository,

  splisoquant --fastq isoquant_tests/simple_data/chr9.4M.ont.sim.fq.gz \
  --reference isoquant_tests/simple_data/chr9.4M.fa.gz \
  --genedb isoquant_tests/simple_data/chr9.4M.gtf.gz \
  --data_type nanopore --complete_genedb -p TEST_DATA --output splisoquant_test
  

• To run Spl-IsoQuant on aligned reads (make sure your BAM is sorted and indexed) use the following command:

  splisoquant --reference /PATH/TO/reference_genome.fasta \
  --genedb /PATH/TO/gene_annotation.gtf \
  --bam /PATH/TO/sample1.sorted.bam /PATH/TO/sample2.sorted.bam \
  --data_type (assembly|pacbio_ccs|nanopore) -o OUTPUT_FOLDER
  

• If using official annotations containing gene and transcript features use --complete_genedb to save time.

• Using reference annotation is optional since version 3.0, you may preform de novo transcript discovery without providing --genedb option:

  splisoquant --reference /PATH/TO/reference_genome.fasta \
  --fastq /PATH/TO/sample1.fastq.gz /PATH/TO/sample2.fastq.gz \
  --data_type (assembly|pacbio_ccs|nanopore) -o OUTPUT_FOLDER
  

• If multiple files are provided, Spl-IsoQuant will create a single output annotation and a single set of gene/transcript expression tables.

• To perform different types of analysis, check out --analysis option.

• For single-cell and spatial transcriptomic data, check out --mode option.