aFC-n calculates the effect size of conditionally independent Expression Quantitative Trait Loci (eQTLs) based on allelic Fold Change (aFC), which could be used to predict genetically driven gene expressionan and allelic imbalance using conditional eQTL data. This script calculates aFCs using least squares optimization (levenberg-marquardt) for a set of eQTLs given a set of gene expressions and phased VCF file. See the manuscript for method description.

First install all the dependencies by doing:

pip3 -r requirements.txt

You will also need gcc and the python 3 development headers. If you are on a Debian based distro, do:

sudo apt install gcc python3-dev

Or on RHEL derivatives:

sudo dnf install gcc python3-devel

Finally, compile the cython files:

bash compile.sh

Running the least squares optimization on several cores is highly recommended, see the plot below as well as the usage examples for more details.

Calculating aFCs without confidence intervals on a single core:

python3 afcn.py --vcf input_vcf.gz --expr input_expressions.gz --eqtl eqtls.txt --output output.txt

Calculating aFCs with confidence intervals on a single core:

python3 afcn.py --conf --vcf input_vcf.gz --expr input_expressions.gz --eqtl eqtls.txt --output output.txt

Calculating aFCs with confidence intervals using 12 cores:

python3 afcn.py -j 12 --conf --vcf input_vcf.gz --expr input_expressions.gz --eqtl eqtls.txt --output output.txt

Calculating aFCs with confidence intervals using 12 cores, with the expressions being in GCT format:

python3 afcn.py --gct -j 12 --conf --vcf input_vcf.gz --expr input_expressions.gz --eqtl eqtls.txt --output output.txt

Calculating aFCs with confidence intervals using 12 cores, with the expressions being log transformed and normalized (--logtransform will do log-transform, --normalize will do log-transform and normalization) :

python3 afcn.py --normalize --logtransform -j 12 --conf --vcf input_vcf.gz --expr input_expressions.gz --eqtl eqtls.txt --output output.txt

--vcf VCF-FILE Genotype VCF

--expr EXPR-FILE Expressions file

--eqtl eQTL-FILE File containing QTL to calculate allelic fold change

--output OUT-FILE Output file name

--nthreads N Number of threads to do fitting on

--conf Calculate confidence intervals for aFC estimates

--normalize Expressions matrix has not been normalized yet

--logtransform Expressions matrix has not been log transformed yet

--splitexpr If set, the individual names in the expressions file will be split on “-” characters and the parts of the name on the two side of the first “-” character will be retained.

--gct If set, it will be assumed that the Expressions file is in gct format

IMPORTANT: The REF and ALT information in the VCF file should match the REF and ALT information in the EQTL matrix

The script expects a gzipped file as an input for gene counts. The input gene counts should be in the format:

Name, sample_id1, sample_id2..

Where Name is a column that has the gene ID (such as ENSG00000224533), which should be in the same format as the gene IDs in the EQTL file.

This file should contain gene IDs, variant IDs that match - it can also contain other stuff, but it needs to contain at least these two columns:

gene_id	variant_id other_stuff1 other_stuff2...

#CHROM  POS     ID      REF     ALT     QUAL    FILTER  INFO  sample_id1  sample_id2...

where #CHROM is the chr # POS is the position, ID is the variant ID in the format chr1_13550_G_A_b38, REF is the reference allele, ALT is the alternative allele. A tabix index needs to be generated for the gzipped vcf file:

bgzip vcf_file.vcf && tabix -p vcf vcf_file.vcf.gz

Your output will be your input EQTL matrix, except that the resulting columns will be added as the additional columns at the end.

log2_aFC - the resulting aFC values

log2_aFC_error - Standard error of the aFCs

log2_aFC_c0 - The residual from the fit

log2_aFC_min_95_interv - The lower 95% conf interval for the aFC value

log2_aFC_plus_95_interv - The upper 95% conf interval for the aFC value

log2_aFC_c0_min_95_interv - The lower 95% conf interval for the residual

log2_aFC_c0_plus_95_interv - The upper 95% conf interval for the residual