Running xipher

Genotyping

TODO: sburger, alecw

Prepare VCF

Load the VCF created in Genotyping step, optionally filter based on gnomAD, and prepare it for xipher.

xipher::prepareVcfClp(outVcfPath, inVcfPath, gnomAdPath)

Prepare digital allele counts (DAC)

1. Run Drop-seq GatherDigitalAlleleCounts on the BAM file for each reaction.

   GatherDigitalAlleleCounts INPUT=your.bam \
    OUTPUT=your.dac.txt.gz \
    VCF=your.vcf \
    CELL_BC_FILE=your.cell_barcodes.txt
    HET_SNPS_ONLY=true \
    SINGLE_VARIANT_READS=true \
    MULTI_GENES_PER_READ=false \
    LOCUS_FUNCTION_LIST=INTRONIC \
    POLYMORPHIC_SNPS_ONLY=false \
    GQ_THRESHOLD=20
   

   • CELL_BC_FILE is a file with the cell barcodes for the reaction.
   • VCF is the file produced as part of genotyping step. TODO: sburger, is this right?
   • INPUT: the BAM file for the reaction.
   • OUTPUT: the output file. It must have extension .dac.txt.gz, and the filename before the extension will be used to prefix the cell barcodes to disambiguate across reactions.
2. Aggregate the DAC files for all reactions into a single file and prepare for xipher.

   xipher::prepareDacClp(outDacPath, dacPaths, annotationsPath, vcfPath)
   

   • dacPaths is a vector of paths to the DAC files created above.
   • annotationsPath is the path to the annotations file created in GTF preparation step.
   • vcfPath is the path to the VCF file created in Prepare VCF step.

Phase X SNPs

xipher::phaseClp(outPhasePath,preparedDacPath)

• preparedDacPath is the file created by xipher::prepareDacClp()

Call X-inactivation

xipher::callActiveXClp(outErrorRatePath, outXCallsPath, preparedDacPath, phasePath)

• preparedDacPath is the file created by `xipher::prepareDacClp()
• phasePath is the file created by xipher::phaseClp()