B6.129S7-Rho^{tm1.1(RHO*/TagRFP-T)Twe}/MrobJ

Stock number: 040788
Product name: P23H-Rho-RFP
Strain synonyms: P23H-hRho-TagRFP-T
Research areas: Mouse/Human Gene Homologs, Neurobiology Research, Research Tools, Sensorineural Research

Description

P23H-Rho-RFP knock-in mice carry the human RHO (rhodopsin) gene with the P23H mutation fused to TagRFP fluorescent reporter in place of the endogenous Rho gene. These mice exhibit progressive retinal degeneration characteristics observed in patients with autosomal dominant retinitis pigmentosa (adRP).

Of note, the donating laboratory has also made their WT-hRho-TagRFP-T knock-in mice (expressing the human wildtype RHO sequence) available as Stock No. 040849.

Detailed description

The P23H-Rho-RFP knock-in mice carry the human RHO (rhodopsin) gene with a nucleotide substitution at codon 23, from CCC to CAC, encoding the amino acid substitution of proline to histidine at position 23, P23H, TagRFP-T, and the C-terminal 9 residues of human rhodopsin in the endogenous mouse Rho locus. The P23H mutation is one of the most common causes of autosomal dominant retinitis pigmentosa (adRP). Homozygous mice are viable and fertile. Both homozygous and heterozygous P23H-Rho-RFP knock-in mice exhibited retinal degradation; however, the homozygous phenotype was more severe and progressed rapidly. (1 cell thickness of ONL by 90 days) Heterozygotes exhibited slower progression, with mutant RHO mislocalization in the inner segments first observed 14 days postnatally. The ONL at 364 days in the heterozygote was 40% of that of WT at 360 days. The presence of RFP enables direct comparison and quantification with the WT-hRho-TagRFP-T knock-in mice (Stock No. 040849) counterpart. The RFP (Tag-RFP-T) fluorescence may be viewed directly or after fixation and co-immunolabeling. These humanized P23H-Rho-RFP mice, which carry a fluorescent tag for quantification, may be useful in studying retinal degeneration diseases, such as autosomal recessive retinitis pigmentosa (adRP), for analyzing the localization of the mutant P23H-Rho protein.

Of note, the donating laboratory has also made their WT-hRho-TagRFP-T knock-in mice (expressing the human wildtype RHO sequence) available as Stock No. 040849.

Development

A targeting vector consists of a human RHO genomic sequence (reference sequence NCBI ID: NG_009115.1) encoding the P23H mutation in the first exon and a fusion to Tag-RFP-T at the C-terminus and an additional C-terminal 9 residues of human rhodopsin (TETSQVAPA, the epitope for the anti-1D4 monoclonal antibody and a putative OS targeting signal), a STOP codon and an endogenous polyA signal followed by lox511-flanked human HPRT1 minigene. An additional 1D4 signaling sequence was added to the carboxy-terminus in case the C-terminal RFP fusion tag may inhibit the endogenous RHO 1D4 sequence. The construct was electroporated into 129S7/SvEvBrd-Hprt1^b-m2 derived AB2.2 embryonic stem (ES) cells to insert 7,465 bp [hRHO (6,706 bp), Tag-RFP-t (732 bp), extra "1D4" sequence (27 bp)] into the mouse Rho locus. Correctly targeted ES cells were injected into recipient mouse blastocysts. The resulting chimeric animals were crossed to C57BL/6 mice to generate germline founders. Founder mice carrying the HPRT-P23H-Rho-RFP allele were crossed to GDF-9-iCre mice on the C57BL/6 genetic background to remove the HPRT1 minigene and screened to ensure germline transmission of the correct targeted sequence without HPRT. P23H-Rho-RFP mice were then extensively backcrossed to C57BL/6 mice by the donating laboratory (see SNP note below) and no longer contain the Cre recombinase transgene. Upon arrival at The Jackson Laboratory, the mice were crossed to C57BL/6J (Stock No. 000664) at least once to establish the colony.

In 2026, a 48 SNP (single nucleotide polymorphism) panel analysis, with 43 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the males sent to The Jackson Laboratory Repository. Three of the 43 markers throughout the genome were segregating with 129 on Chromosome 6 and Chromosome 12. The segregation on Chromosome 6 is likely a passenger segment from Rho mutation.

Allele

Symbol: Rho^{tm1.1(RHO*/TagRFP-T)Twe}
Name: targeted mutation 1.1, Theodore G Wensel
MGI accession ID: MGI:8221592
Generation method: Targeted
Attributes: Reporter; Inserted expressed sequence; Humanized sequence
Synonyms: P23HhRhoRFP; P23H-hRho-TagRFP; P23H-Rho-RFP
Marker symbol: Rho
Marker name: rhodopsin
Marker synonyms: CSNBAD1; L opsin; Long Wavelength Sensitive opsin; LWS opsin; MGC:21585; MGC:25387; Noerg1; Opn2; Ops; opsin 2; Red Opsin; Rod Opsin; RP4
Chromosome: 6
Strain of origin: 129S7/SvEvBrd-Hprt1^b-m2
Molecular note: The endogenous gene was replaced with a human RHO gene with a P23H mutation, a C terminal fusion to the fluorescent Tag-RFP-T and a lox511-flanked HPRT minigene via homologous recombination. An additional 1D4 signaling sequence was added to the carboxy-terminus in case the C-terminal RFP fusion tag may inhibit the endogenous Rho 1D4 sequence. Cre mediated recombination removed the HPRT minigene.