Germline Transformation of the Oriental Fruit Fly Bactrocera dorsalis (Hendel) Using a piggyBac-derived Vector

利用piggyBac轉基因子進行東方果實蠅之生殖細胞系基因轉型

陳秀玲(Shiu-Ling Chen) ； 張誠(Cheng Chang) ； 黃紹毅(Shaw-Yhi Hwang) ； 路光暉(Kuang-Hui Lu)

《台灣昆蟲》 30卷1期 (2010/03) Pp. 27-40

https://doi.org/10.6661/TESFE.2010002

摘要

東方果實蠅(Bactrocera dorsalis (Hendel))為亞洲國家及台灣的重要果樹害蟲。本研究，我們利用生殖細胞系轉型(germline transformation)技術建立轉基因東方果實蠅(transgenic orient fruit fly)，期以進一步發展利用昆蟲轉基因技術(insect transgenic technology)建立害蟲防治的方法。進行基因轉殖所使用的轉基因子(transposon) 為piggyBac；將帶有果蠅泛素啟動子(Drosophila polyubiquitin promoter)調控紅色螢光蛋白基因(DsRed gene)之piggyBac轉基因子載體，以微注射(microinjection)注入胚胎內進行細胞轉型作用，使外來基因嵌入果實蠅基因組(genome)上。試驗結果顯示轉基因之平均成功率為5.3%，此與其他同樣以piggyBac轉基因子轉殖雙翅目昆蟲之結果相近。轉基因東方果實蠅個體均可於各發育期穩定表現紅色螢光蛋白(DsRed)。轉位分析(transposition assay)證實標的片段嵌入基因組的TTAA位置，符合piggyBac轉因子之作用特性；而南方墨點法(Southern blot)則顯示該標的片段可穩定遺傳至子代，目前已超過65代。

關鍵字

生殖細胞系轉型 ； 基因轉殖 ； piggyBac轉基因子 ； 微注射 ； 東方果實蠅

並列摘要

The oriental fruit fly, Bactrocera dorsalis (Hendel), is a serious pest of fruit crops in most Asian countries including Taiwan. In this study, we have developed a stable germline transformation system with the ultimate goal of using transgenic technology to control this insect pest. A non-drosophilid transposable element, i.e. piggyBac, which contains the red fluorescent reporter gene, DsRed, under control of the Drosophila polyubiquitin promoter, was used to transform B. dorsalis. The average transformation frequency with different ratios of reporter and helper vectors was 5.3%, which is comparable to those reported in other piggyBac-transformed flies. The red fluorescence of DsRed is expressed in all developmental stages of the transgenic B. dorsalis. Transposition assays show that the exogenous fragment is integrated into TTAA sites. In addition, Southern blot analyses verify that the exogenous genes are stably inherited over 65 generations.

並列關鍵字

germline transformation ； transgenesis ； piggyBac ； microinjection ； Bactrocera dorsalis