Combining Results

After you have run each of the Demultiplexing and Doublet Detecting softwares you would like, it is helpful to convert them to similar nomenclarture and combine the results into a single dataframe. In addition, we have found it helpful to generate summaries of each of the combinations of softwares identified. To help streamline this process, we have provided a script that will easily integrate all the softwares you have run into a single dataframe and to generate a summary file for all the software combinations and if you ran demultiplexing softwares, it will also generate a demultiplexing summary file for the individual and cluster assignments from the demultiplexing softwares.

Data

In order to use our script to combine the results from the various demultiplexing and doublet detecting softwares, you need the following:

Required

• Output directory ($OUTDIR)

• Path to results of each of the softwares you would like to merge into a single dataframe.

  • You need to provide the path to at least one software result, otherwise, it will not run.

Merging Results with Combine_Results.r

The script has multiple options to provide the paths to each of the software results you would like to run. To see each of the options, simply run:

singularity exec Demuxafy.sif Combine_Results.R -h

Providing the possible parameter options:

usage: Combine_Results.R
     [-h] -o OUT [-d DEMUXLET] [-f FREEMUXLET] [-g FREEMUXLET_ASSIGNMENTS]
     [-s SCSPLIT] [-w SCSPLIT_ASSIGNMENTS] [-u SOUPORCELL]
     [-x SOUPORCELL_ASSIGNMENTS] [-v VIREO] [-e DOUBLETDECON]
     [-t DOUBLETDETECTION] [-i DOUBLETFINDER] [-n SCDBLFINDER] [-c SCDS]
     [-r SCRUBLET] [-l SOLO]

optional arguments:
  -h, --help            show this help message and exit
  -o OUT, --out OUT     The file where results will be saved
  -d DEMUXLET, --demuxlet DEMUXLET
                        Path to demuxlet results. Only use this option if you
                        want to include the demuxlet results.
  -f FREEMUXLET, --freemuxlet FREEMUXLET
                        Path to freemuxlet results. Only use this option if
                        you want to include the freemuxlet results.
  -g FREEMUXLET_ASSIGNMENTS, --freemuxlet_assignments FREEMUXLET_ASSIGNMENTS
                        Path to freemuxlet cluster-to-individual assignments.
                        Only use this option if have used reference SNP
                        genotypes to assign individuals to clusters for the
                        freemuxlet results.
  -s SCSPLIT, --scSplit SCSPLIT
                        Path to scSplit results. Only use this option if you
                        want to include the scSplit results.
  -w SCSPLIT_ASSIGNMENTS, --scSplit_assignments SCSPLIT_ASSIGNMENTS
                        Path to scSplit cluster-to-individual assignments.
                        Only use this option if you have used reference SNP
                        genotypes to assign individuals to clusters for the
                        scSplit results.
  -u SOUPORCELL, --souporcell SOUPORCELL
                        Path to souporcell results. Only use this option if
                        you want to include the souporcell results.
  -x SOUPORCELL_ASSIGNMENTS, --souporcell_assignments SOUPORCELL_ASSIGNMENTS
                        Path to souporcell cluster-to-individual assignments.
                        Only use this option if you have used reference SNP
                        genotypes to assign individuals to clusters for the
                        souporcell results.
  -v VIREO, --vireo VIREO
                        Path to vireo results. Only use this option if you
                        want to include the vireo results.
  -e DOUBLETDECON, --DoubletDecon DOUBLETDECON
                        Path to DoubletDecon results. Only use this option if
                        you want to include the DoubletDecon results.
  -t DOUBLETDETECTION, --DoubletDetection DOUBLETDETECTION
                        Path to DoubletDetection results. Only use this option
                        if you want to include the DoubletDetection results.
  -i DOUBLETFINDER, --DoubletFinder DOUBLETFINDER
                        Path to DoubletFinder results. Only use this option if
                        you want to include the DoubletFinder results.
  -n SCDBLFINDER, --scDblFinder SCDBLFINDER
                        Path to scDblFinder results. Only use this option if
                        you want to include the scDblFinder results.
  -c SCDS, --scds SCDS  Path to scds results. Only use this option if you want
                        to include the scds results.
  -r SCRUBLET, --scrublet SCRUBLET
                        Path to scrublet results. Only use this option if you
                        want to include the scrublet results.
  -l SOLO, --solo SOLO  Path to solo results. Only use this option if you want
                        to include the solo results.

An example command that combines Demuxlet results, Souporcell results, Solo results and Scds results would look like this:

singularity exec Demuxafy.sif Combine_Results.R \
  -o $OUTDIR/combined_results.tsv \
  --demuxlet $DEMUXLET_OUTDIR \
  --souporcell $DEMUXLET_OUTDIR \
  --solo $SOLO_OUTDIR \
  --scds $SCDS_OUTDIR \

Note

The path to the directories will work if the file names are the expected file names. However, if you used a different file naming convention or changed the names, you can also provide the full path to the exact file for each software.

Results and Interpretation

After running the Combine_Results.R script, you should have three (or two if you didn’t have any demultiplexing softwares)

.
├── combined_results_demultiplexing_summary.tsv
├── combined_results_summary.tsv
└── combined_results.tsv

Note

You will only have the combined_results_demultiplexing_summary.tsv file if you included demultiplexing softwares.

Citation

If you used this workflow for analysis, please reference our paper (REFERENCE).