PT - JOURNAL ARTICLE AU - Anny Devoy AU - Georgia Price AU - Francesca De Giorgio AU - Rosie Bunton-Stasyshyn AU - David Thompson AU - Samanta Gasco AU - Alasdair Allan AU - Gemma F. Codner AU - Remya R. Nair AU - Charlotte Tibbit AU - Ross McLeod AU - Zeinab Ali AU - Judith Noda AU - Alessandro Marrero-Gagliardi AU - José M Brito-Armas AU - Michelle Simon AU - Edward O’Neill AU - Jackie Harrison AU - Gemma Atkins AU - Silvia Corrochano AU - Michelle Stewart AU - Lydia Teboul AU - Abraham Acevedo-Arozena AU - Elizabeth M.C Fisher AU - Thomas J. Cunningham TI - Generation, quality control, and analysis of the first genomically humanised knock-in mice for the ALS/FTD genes <em>SOD1, TARDBP</em> (TDP-43), and <em>FUS</em> AID - 10.1101/2021.07.05.451113 DP - 2021 Jan 01 TA - bioRxiv PG - 2021.07.05.451113 4099 - http://biorxiv.org/content/early/2021/07/05/2021.07.05.451113.short 4100 - http://biorxiv.org/content/early/2021/07/05/2021.07.05.451113.full AB - Amyotrophic lateral sclerosis - frontotemporal dementia spectrum disorder (ALS/FTD) is a complex neurodegenerative disease; up to 10% of cases are familial, usually arising from single dominant mutations in &gt;30 causative genes. Transgenic mouse models that overexpress human ALS/FTD causative genes have been the preferred organism for in vivo modelling. However, while conferring human protein biochemistry, these overexpression models are not ideal for dosage-sensitive proteins such as TDP-43 or FUS.We have created three next-generation genomically humanised knock-in mouse models for ALS/FTD research, by replacing the entire mouse coding region of Sod1, Tardbp (TDP-43) and Fus, with their human orthologues to preserve human protein biochemistry, with exons and introns intact to enable future modelling of coding or non-coding mutations and variants and to preserve human splice variants. In generating these mice, we have established a new-standard of quality control: we demonstrate the utility of indirect capture for enrichment of a region of interest followed by Oxford Nanopore sequencing for robustly characterising large knock-in alleles. This approach confirmed that targeting occurred at the correct locus and to map homologous recombination events. Furthermore, extensive expression data from the three lines shows that homozygous humanised animals only express human protein, at endogenous levels. Characterisation of humanised FUS animals showed that they are phenotypically normal compared to wildtype littermates throughout their lifespan.These humanised mouse strains are critically needed for preclinical assessment of interventions, such as antisense oligonucleotides (ASOs), to modulate expression levels in patients, and will serve as templates for the addition of human ALS/FTD mutations to dissect disease pathomechanisms.Competing Interest StatementThe authors have declared no competing interest.