Document Type


Publication Date




JAX Source

Genome Biol 2020 Jul 9; 21:168





CA034196,OD27052,OD026440,AI132963,Director's Innovation Fund.


BACKGROUND: Gene disruption in mouse embryonic stem cells or zygotes is a conventional genetics approach to identify gene function in vivo. However, because different gene disruption strategies use different mechanisms to disrupt genes, the strategies can result in diverse phenotypes in the resulting mouse model. To determine whether different gene disruption strategies affect the phenotype of resulting mutant mice, we characterized Rhbdf1 mouse mutant strains generated by three commonly used strategies-definitive-null, targeted knockout (KO)-first, and CRISPR/Cas9.

RESULTS: We find that Rhbdf1 responds differently to distinct KO strategies, for example, by skipping exons and reinitiating translation to potentially yield gain-of-function alleles rather than the expected null or severe hypomorphic alleles. Our analysis also revealed that at least 4% of mice generated using the KO-first strategy show conflicting phenotypes.

CONCLUSIONS: Exon skipping is a widespread phenomenon occurring across the genome. These findings have significant implications for the application of genome editing in both basic research and clinical practice.


We gratefully acknowledge the contributions of the JAX core for expert assistance with the work described in this publication, especially Mary Barter, Chrystal Snow, Christina Petros, and Heidi Munger at the Genome Technologies, Peter Kutny and Bill Buaas at Genetic Engineering Technologies, Gregg TeHennepe at Computational Sciences, and Nick Gott and Elaine Bechtel at Histopathology and Microscopy Sciences. We thank Dr. Carl P. Blobel for providing frozen spleens from the Rhbdf1v2/v2 mice and for critically reading the manuscript. We thank the members of the Rosenthal laboratory (Drs. Mauro Costa and Milena Furtado) at JAX for technical support and for helpful suggestions. We thank Michelle L. Farley, Cindy Avery, and Todd Nason for maintenance of mouse colonies and for monitoring of early pup survival, and the “eyelids open at birth” phenotype. We thank Lihong Zhao, Rosalinda A. Doty, Richard S. Maser, and Ryuta Ishimura for helpful discussion, and Stephen B. Sampson for critical reading of the manuscript. We also thank Zoe Reifsnyder in preparing illustrations for the manuscript.

This article is licensed under a Creative Commons Attribution 4.0 International License.