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Sci Rep. 2019 Aug 14;9(1):11828. doi: 10.1038/s41598-019-48105-4.

A BAC Transgene Expressing Human CFTR under Control of Its Regulatory Elements Rescues Cftr Knockout Mice.

Author information

1
Dalton Cardiovascular Research Center, University of Missouri, 134 Research Park Dr, Columbia, Missouri, 65211-3300, USA.
2
Department of Biomedical Sciences, University of Missouri, E102 Veterinary Medicine Bldg., Columbia, Missouri, 65211, USA.
3
Departments of Pediatrics, Case Western Reserve University School of Medicine, Cleveland, Ohio, USA.
4
Departments of Genetics and Genome Sciences, Case Western Reserve University School of Medicine, Cleveland, Ohio, USA.
5
Departments of Physiology and Biophysics, Case Western Reserve University School of Medicine, Cleveland, Ohio, USA.
6
Human Molecular Genetics Program, Lurie Children's Research Center, Northwestern University Feinberg School of Medicine, Chicago, IL, 60614, USA.
7
RNA Bioscience Initiative, University of Colorado School of Medicine, Aurora, CO, USA.
8
Department of Histology and Embryology, School of Medicine, University of Athens, Athens, Greece.
9
InterRayBio, LLC, Baltimore, MD, USA.
10
Dalton Cardiovascular Research Center, University of Missouri, 134 Research Park Dr, Columbia, Missouri, 65211-3300, USA. clarkel@missouri.edu.
11
Department of Biomedical Sciences, University of Missouri, E102 Veterinary Medicine Bldg., Columbia, Missouri, 65211, USA. clarkel@missouri.edu.

Abstract

Small-molecule modulators of cystic fibrosis transmembrane conductance regulator (CFTR) biology show promise in the treatment of cystic fibrosis (CF). A Cftr knockout (Cftr KO) mouse expressing mutants of human CFTR would advance in vivo testing of new modulators. A bacterial artificial chromosome (BAC) carrying the complete hCFTR gene including regulatory elements within 40.1 kb of DNA 5' and 25 kb of DNA 3' to the gene was used to generate founder mice expressing hCFTR. Whole genome sequencing indicated a single integration site on mouse chromosome 8 (8qB2) with ~6 gene copies. hCFTR+ offspring were bred to murine Cftr KO mice, producing hCFTR+/mCftr- (H+/m-) mice, which had normal survival, growth and goblet cell function as compared to wild-type (WT) mice. Expression studies showed hCFTR protein and transcripts in tissues typically expressing mCftr. Functionally, nasal potential difference and large intestinal short-circuit (Isc) responses to cAMP stimulation were similar in magnitude to WT mice, whereas small intestinal cAMP ΔIsc responses were reduced. A BAC transgenic mouse with functional hCFTR under control of its regulatory elements has been developed to enable the generation of mouse models of hCFTR mutations by gene editing for in vivo testing of new CF therapies.

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