A versatile vector set for animal transgenesis
A versatile vector set for animal transgenesis
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DOI：
10.1007/s004270000110
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发表时间：
2000-12-01
影响因子：
2.4
通讯作者：
Wimmer, EA
中科院分区：
文献类型：
--
作者：
Horn, C;Wimmer, EA
为了更好地理解发育和进化过程，对一系列不同节肢动物的遗传操作是一个非常理想的目标。到目前为止，一个主要的障碍是难以获得标记基因，这些标记基因可以方便可靠地鉴定转基因动物。在这里，我们提出了一个基于混杂转座子mariner， Hermes和piggyBac的生殖系转化的多功能载体集。在这些载体中，我们引入了一个潜在的通用标记系统，该系统由含有三个Pax-6同型二聚体结合位点的人工启动子组成。该启动子驱动增强绿色荧光蛋白（EGFP）光谱变体在幼虫、蛹和成虫的光感受器中的强烈表达。使用特殊的过滤集，黄色（EYFP）和青色（ECFP）变体是完全可区分的，因此代表了一对可分离的标记。此外，我们采用了一种简单的基于质粒的转座试验系统，以便在将我们的载体用于更费力的种系转化实验之前，在不同的节肢动物物种中进行快速功能测试。利用这个系统，我们证明了我们的载体在黑腹果蝇和雄果蝇中都能转置。
Genetic manipulation of a series of diverged arthropods is a highly desirable goal for a better understanding of developmental and evolutionary processes. A major obstacle so far has been the difficulty in obtaining marker genes that allow easy and reliable identification of transgenic animals. Here, we present a versatile vector set for germline transformation based on the promiscuous transposons mariner, Hermes and piggyBac. Into these vectors, we introduced a potentially universal marker system that is comprised of an artificial promoter containing three Pax-6 homodimer binding sites. This promoter drives strong expression of spectral variants of the enhanced green fluorescent protein (EGFP) in larval, pupal, and adult photoreceptors. Using special filter sets, the yellow (EYFP) and cyan (ECFP) variant are fully distinguishable and therefore represent a separable pair of markers. Furthermore, we adapted a simple plasmid-based transposition assay system to enable quick functional tests of our vectors in different arthropod species before employing them in more laborious germline transformation experiments. Using this system we demonstrate that our vectors transpose in both Drosophila melanogaster and Drosophila virilis.