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X. Zhang, H. Tseng; Creating a Conditional–Null Mouse Model for Basonuclin . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1589.
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© ARVO (1962-2015); The Authors (2016-present)
Basonuclin is a zinc finger protein mainly present in stratified epithelia such as corneal epithelium. To investigate basonuclin function in this tissue, we constructed a conditional–null mouse model.
The exon 4 of basonuclin gene, which encodes the critical zinc fingers and the nuclear localization signal, was targeted for excision by the Cre recombinase. The targeting vector, which was constructed by ET recombination with a 12.7kb genomic sequence of the basonuclin gene, was introduced into 129s/R1 ES cells by electroporation. Homologous recombination was detected by Southern hybridization. Two independently isolated Bfl–neo/+ clones were injected into C57BL/6 blastocysts at the Transgenic & Chimeric Mouse Facility at the University of Pennsylvania. Southern hybridization and PCR were used to confirm the genotype.
The targeting vector was constructed by flanking exon 4 of the basonuclin gene with two lox P sites. After electroporating the targeting vector into ES cells, 224 G418–resistent colonies were isolated, of which, 7 were shown to contain the correctly recombined targeting sequence by Southern hybridization with 5' and 3' external probes and an internal probe. Four male chimeras with 100% agouti coat color transmitted the transgene to their progeny. Both transgenic heterozygotes (Bfl–neo/+) and homozygotes (Bfl–neo/fl–neo) are normal and fertile. By breeding the Bfl–neo/+ mice with ZP–3/Cre mice, we obtained B+/– mice, which is also apparently normal. We plan to mate the Bfl–neo/fl–neo mice with K12–IRES/Cre mice to create a corneal epithelial specific basonuclin knock–out mouse.
We have succeeded in generating a basonuclin conditional–null mouse model for corneal epithelium. This mouse model is valuable to reveal the role of basonuclin in corneal epithelium development and maintenance.
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