Biallelic genome engineering to create isogenic induced pluripotent stem cells modelling Huntington's disease.

Abstract

We developed Huntington's disease (HD) modelling induced pluripotent stem cells (iPSCs) by genome engineering of iPSCs from healthy donors. For this, we established a homologous-recombination-based biallelic substitution technique called the allele-specific universal knock-in system (asUKiS). asUKiS allows for scarless and allele-by-allele substitution of the entire region encompassing not only the polyQ-repeat but also the associated genetic modifiers surrounding the repeat region, allowing us to generate five iPSC lines with identical genetic modifiers on both alleles, differing only in polyQ repeat numbers. All cell lines were validated by allele-specific genotyping to confirm the precise engineering of both alleles. Even for modelling autosomal dominant diseases, our approach of employing biallelic modification may offer the distinct advantage enabling the investigation of the effects of specific genomic mutations with minimal interference from genetic background noise.