Development of transposon-vector-systems in insects.

在分析模式生物的基因功能时，转化技术是最重要的，例如，用野生型基因或修饰基因转化突变体。在昆虫中，除了果蝇外，还没有开发出有效的转化方法。利用转座元件，P-元件可以高效地转化果蝇。这种方法的另一个优点是大多数转基因都是以单拷贝插入的，便于分析给定的基因功能。本项目的目的是开发利用转座元件的转化系统。本研究使用的材料是：黑腹果蝇、蚊子(白纹伊蚊)和家蚕(Bombyx Mori)。有些种类的蚊子被认为是传播疾病的媒介，例如疟疾、登革热。蚕虫是一种有用的昆虫，不仅用于纺织品，而且还用于从丝素腺体中生产生物分子。因此，在这些物种中需要一种高效的转化体系。Electrporat…更多的离子方法现在被广泛用于将生物从细菌转化为哺乳动物。我们试图将这种方法应用于使用果蝇胚胎的昆虫。在以克隆的α-淀粉酶基因为报告基因的初步实验中，检测到3龄幼虫的淀粉酶活性。然而，使用P元件载体和辅助子，从3000个胚胎中没有获得转化子。为了将眼色基因作为转化标记，用甲基磺酸乙酯(EMS)诱导白纹伊蚊眼色突变。到目前为止，已经确定了两个突变。一个是位于性染色体上的棕色眼1，花药是位于常染色体上的棕色眼2。Bm1是家蚕的一种正弦分子。我们已经测定了15个完整的Bm1.1元件的完整序列。序列比较和系统发育树表明，它们被划分为三个组。这些结果表明，Bm1起源于树木的祖先元素，并通过逆转录分散在基因组中。较少

Transformation techniques are most important to analyze gene functions in model organisms, e. g. a mutant is transformed with a wild-type gene or a modified one. In insects, effective method for transformation had not been developed except for Drosophila. Using a transposable element, P-element, Drosophila can be transformed with high efficiency. Another advantage of this method is that tha most transgene is inserted with single copy and it eases to analyze given gene functions.The aim of this project is to develop transformation system using transposable elements. The materials used in this study are : Drosophila melanogaster, a mosquito species (Aedes albopictus) and the silkworm (Bombyx mori). Some species of mosquito are known as vectors for diseases, e. g. malaria, dengue. The silk worm is a useful insect not only for the textiles but also for the production of biomolecules from fibroin-gland. Therefore, an effetive transformation systems were desired in these species.Electroporat … More ion method is now widely used to transform organisms from bacteria to mammals. We tried to aply this method to insects using Drosophila embryos. In preliminary experiment with a cloned alpha-amylase gene as a reporter gene, the amylase activity was detected in the third instar larvae. Using P-element vector and a helper, however, no transformant was obtained out of 3000 embryos.To apply an eye color gene as a transformation marker, eye color mutations of Aedes albopictus were induced by Ethyl methan sulfonate (EMS). Until now, two mutations were identified. One is brown eye1 located on the sex-chromosome and anther is brown eye2 located on an autosome.Bm1 is a kind of SINE element of Bombyx mori. We had determined the complete sequences of fifteen of complete Bm1.1 elements. Sequence comparisons and a phylogenetic tree showed that they are classified into three groups. These results suggest that Bm1 was originated from tree ancestral elements and dispersed in the genome by reverse-transcription. Less