Tracing retinal fiber trajectory with a method of transposon-mediated genomic integration in chick embryo
Tracing retinal fiber trajectory with a method of transposon-mediated genomic integration in chick embryo
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DOI：
10.1111/j.1440-169x.2008.01067.x
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发表时间：
2008-10-01
影响因子：
2.5
通讯作者：
Nakamura, Harukazu
中科院分区：
文献类型：
--
作者：
Harada, Hidekiyo;Takahashi, Yoshiko;Nakamura, Harukazu
我们报告方便的视网膜纤维示踪剂通过卵内电穿孔的示踪剂cDNA。示踪蛋白如绿色荧光蛋白的长期和稳定表达通过示踪蛋白表达盒的转座子介导的基因组整合来实现。我们进行了共电穿孔的质粒含有CAGGS-示踪cDNA侧翼的Tol 2转座因子沿着与转座酶表达载体的视泡的鸡胚在第11阶段。通过选择电极，我们可以标记一大群视网膜神经节细胞，或一小群视网膜神经节细胞;平行电极确保大面积视网膜的转染，针型电极标记视网膜的小区域。在视顶盖的对侧，视网膜纤维轨迹和终末带（TZ）可以被精确的视网膜定位方式检测到。该方法的优点在于，我们可以显示视网膜纤维轨迹与负责同一标本中视网膜纤维寻路的分子的关系。
We report convenient retinal fiber tracing by transfecting the tracer cDNA by in ovo electroporation. Long-term and stable expression of tracer proteins such as green fluorescent protein is achieved by transposon-mediated genome integration of the tracer protein expression cassette. We carried out coelectroporation of a plasmid containing CAGGS-tracer cDNA flanked by the Tol2 transposable element along with a transposase expression vector to the optic vesicle of chick embryos at stage 11. By selecting electrodes, we can label a large group of retinal ganglion cells, or a small group of retinal ganglion cells; parallel electrodes assure transfection of large areas of the retina, and needle type electrodes label small areas of the retina. The retinal fiber trajectory and terminal zone (TZ) could be detected in the precise retinotopic manner on the contra-lateral side of the optic tectum. The method has advantage in that we can show the retinal fiber trajectory in relation to the molecules that are responsible for pathfinding for the retinal fibers in the same specimen.