Cre–Transgenic Mouse Line for Conditional Gene Knockout in Retinal Rod Bipolar Cells

Abstract

Abstract: : Purpose: Some progressive retinal diseases are caused by degeneration of neurons in the rod–pathway. It remains unknown how rod bipolar cells contribute to retinal degenerative disease. This study was conducted to establish a transgenic mouse line expressing Cre recombinase specifically in retinal rod bipolar cells. Methods: IRES–Cre–cDNA fragment was inserted into a 173–kb bacterial artificial chromosome (BAC) carrying the intact Pcp2 gene using Red–mediated recombineering. Transgenic mice were generated with the modified BAC and identified by Southern blot analysis and PCR. The activity of Cre–recombinase was analyzed in progenies from Cre–transgenic mice crossed with ROSA26 or Z/EG reporter mice. Results: The genotypes of BAC–Pcp2–IRES–Cre transgenic mice were confirmed by PCR with Cre–specific primers. X–gal staining showed that the activities of Cre–recombinase were present in cerebellar Purkinje cells and retinal inner nuclear layer, where rod bipolar cells are located. Immunofluorescent double–staining with anti–GFP antibody and anti–PKCα (specific marker for retinal rod bipolar cells) antibody revealed that Cre–recombinase activity localized exclusively to the rod bipolar cells in the retina. Conclusions: A transgenic mouse line BAC–Pcp2–IRES–Cre that expresses Cre–recombinase in retinal rod bipolar cells has been successfully established. It will be a useful new tool for investigating the effects of retinal rod bipolar cell–specific gene inactivation and invaluable for future studies on retinal functions.

Keywords: bipolar cells • retinal development • gene transfer/gene therapy