An expression-based variant impact phenotyping protocol to predict the impact of gene variants in cell lines.

We describe a bioinformatics protocol for eVIP2 (expression-based variant impact phenotyping). eVIP2 can predict a gene variant's functional impact by comparing gene expression signatures induced by introduction of wild-type versus mutant cDNAs in cell lines. The predicted functional outcomes of the variants include gain-of-function, loss-of-function, change-of-function, or neutral. eVIP2 improves upon eVIP by being applicable to RNA-seq data and providing pathway-level functional predictions for each mutation. Here, we detail how to run eVIP2 on RNA-seq data from two RNF43 variants. For complete details on the use and execution of this protocol, please refer to Thornton et al. (2021).