Rhox-c floxed mice have loxP sites flanking all 33 genes of the Rhox cluster. Exposure to Cre recombinase removes the floxed sequence - creating a null allele. These Rhox-c floxed mice may be useful in studying spermatogenesis and reproductive biology.
Dr. Miles Wilkinson, University of California San Diego
The X-linked reproductive homeobox cluster (Rhox, also known as Rhox-c) encodes a group of 33 genes, all of which encode homeobox proteins that are primarily expressed in the reproductive tract.
The Rhox-c-floxed allele has loxP sites flanking all 33 genes of the Rhox gene cluster. Exposure to Cre recombinase removes the floxed sequence - creating a null allele. Prior to introduction of Cre recombinase, heterozygous females, hemizygous males, or females homozygous for the Rhox-c-floxed allele are viable and fertile with normal breeding, and have no gross physical or behavioral abnormalities.
When Rhox-c-floxed mice are bred to mice that express Cre recombinase, the resulting offspring will have the entire Rhox locus deleted - creating a null allele in the cells/tissues as determined by the Cre promoter.
For example, when bred to germline Cre-expressing mice (E2A-Cre; see Stock No. 003724), the resulting Rhox-c-KO global knock-out hemizygotes were confirmed to have no expression of any of the Rhox genes tested in testes homogenates (by qRT-PCR). Those Rhox-c-KO global hemizygous knock-out males exhibit increased embryonic lethality, while surviving pups are runted when on a mixed genetic background including contributions from C57BL/6, FVB/NJ, and 129. The incidence of embryonic lethality appears to increase concomitantly with the contribution of C57BL/6. Surviving homozygotes on the mixed background exhibit reduced testes size which fail to grow during sexual maturity, as well as reduced sperm counts and vacuolar seminiferous tubules lacking in germ cells.
The Rhox-c floxed allele was created in the laboratory of Dr. Miles F. Wilkinson (University of California, San Diego). To generate Rhox-c floxed mice, two targeting vectors were designed to insert loxP sites at the 5’ and 3’ ends of the X-linked Rhox homeobox gene cluster. The first vector contained two loxP sites surrounding a neomycin resistance cassette and two homology arms matching the upstream intergenic region of Rhox1. The second vector contained a loxP-flanked puromycin resistant cassette and two homologous arms for recombination with the second intronic region of Rhox13. The vectors were sequentially electroporated into 129-derived ES cells. Correctly targeted ES cells were injected into recipient blastocysts and the resulting chimeric mice were crossed to C57BL/6J for germline transmission. The line was then crossed to EIIa-cre to remove the neo and puromycin cassettes, leaving a single loxP site at each end of the Rhox cluster. The Rhox-c-floxed line was then crossed again to C57BL/6J to remove the EIIa-cre allele, and subsequently backcrossed to C57BL6/J at least 6 additional generations prior to sending males to The Jackson Laboratory Repository in 2020. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664).
|Allele Name||targeted mutation 1, Miles F Wilkinson|
|Allele Type||Targeted (Conditional ready (e.g. floxed))|
|Allele Synonym(s)||Rhox-cfl; Rhox-c-floxed|
|Gene Symbol and Name||Rhox, reproductive homeobox cluster|
|Strain of Origin||129|
|Molecular Note||A loxP sites were inserted upstream of Rhox1 and into the second intron of Rhox13.|
Rhox is X-linked. Both heterozygous males and homozygous females are viable and fertile with normal breeding, and no reported gross phenotypic or behavioral abnormalities.
When maintaining a live colony, heterozygous mice may be bred together, to wildtype mice from the colony or to C57BL/6J inbred mice (Stock No. 000664).
Alternatively, homozygous females may be bred to heterozygous males.
When using the Rhox-c flox
mouse strain in a publication, please cite the originating article(s) and include JAX stock #035194 in your Materials and Methods section.