Zhang Xin-Mei, Chen Bai-Yu, Ng Alam Hoi-Lam, Tanner Julian A, Tay David, So Kwok-Fai, Rachel Rivka A, Copeland Neal G, Jenkins Nancy A, Huang Jian-Dong
Department of Biochemistry, University of Hong Kong, SAR, China.
Invest Ophthalmol Vis Sci. 2005 Oct;46(10):3515-20. doi: 10.1167/iovs.04-1201.
To establish a transgenic mouse line that expresses Cre-recombinase in retinal rod bipolar cells for the generation of rod bipolar cell-specific knockout mutants.
The IRES-Cre-cDNA fragment was inserted into a 173-kb bacterial artificial chromosome (BAC) carrying the intact Pcp2 gene, by using red-mediated recombineering. Transgenic mice were generated with the modified BAC and identified. The Cre-transgenic mice were crossed with ROSA26 and Z/EG reporter mice to detect Cre-recombinase activity.
X-gal staining showed that strong Cre-recombinase activities were present in retinal inner nuclear layers and cerebellar Purkinje cells. Double staining with an anti-GFP antibody and an anti-PKCalpha antibody (specific for retinal rod bipolar cells) revealed that Cre-recombinase activity localized exclusively to the rod bipolar cells in the retina.
A mouse BAC-Pcp2-IRES-Cre transgenic line that expresses Cre-recombinase in retinal rod bipolar neurons has been established. Because mutations in some ubiquitously expressed genes may result in retinal degenerative diseases, the mouse strain BAC-Pcp2-IRES-Cre will be a useful new tool for investigating the effects of retinal rod bipolar cell-specific gene inactivation.
建立一种在视网膜视杆双极细胞中表达Cre重组酶的转基因小鼠品系，用于生成视杆双极细胞特异性敲除突变体。
通过红介导的重组工程，将IRES-Cre-cDNA片段插入携带完整Pcp2基因的173 kb细菌人工染色体（BAC）中。用修饰后的BAC生成转基因小鼠并进行鉴定。将Cre转基因小鼠与ROSA26和Z/EG报告基因小鼠杂交，以检测Cre重组酶活性。
X-gal染色显示，视网膜内核层和小脑浦肯野细胞中存在较强的Cre重组酶活性。用抗GFP抗体和抗PKCalpha抗体（对视杆双极细胞特异）进行双重染色显示，Cre重组酶活性仅定位于视网膜中的视杆双极细胞。
已建立了在视网膜视杆双极神经元中表达Cre重组酶的小鼠BAC-Pcp2-IRES-Cre转基因品系。由于一些普遍表达的基因发生突变可能导致视网膜退行性疾病，BAC-Pcp2-IRES-Cre小鼠品系将成为研究视网膜视杆双极细胞特异性基因失活效应的一种有用的新工具。
