Establishment of a stable transgenic strain in a pelagic egg spawning marine teleost, Nibe croaker Nibea mitsukurii
Establishment of a stable transgenic strain in a pelagic egg spawning marine teleost, Nibe croaker Nibea mitsukurii
复制标题
DOI：
10.1016/j.aquaculture.2011.01.042
复制
发表时间：
2011-03
期刊：
影响因子：
4.5
通讯作者：
Yoji Yamamoto;N. Kabeya;Y. Takeuchi;K. Higuchi;T. Yatabe;K. Tsunemoto;R. Yazawa;Takaichi Kawamura;G. Yoshizaki
中科院分区：
文献类型：
--
作者：
Yoji Yamamoto;N. Kabeya;Y. Takeuchi;K. Higuchi;T. Yatabe;K. Tsunemoto;R. Yazawa;Takaichi Kawamura;G. Yoshizaki
转基因是全球粮食生产的一项重要技术。转基因鱼种可以通过显微注射建立，但这种有效的方法还没有在海洋鱼类上开发，因为处理它们的小而脆弱的远洋鱼卵和幼体存在困难。本论文报道了一种转基因大黄鱼Nibea mitsukurii的生产。Nibea mitsukurii是一种产小型远洋鱼卵的海鱼。采用显微注射技术，将虹鱼热休克相关基因71(Hsc71)启动子/增强子驱动的增强型绿色荧光蛋白(EGFP)基因直接注入受精卵。在610枚显微注射的卵中，498枚(81.6%)存活到14体期(受精后24小时)，其中426枚(69.8%)表现出高嵌合的GFP瞬时表达，并被选择饲养至性成熟。在存活的鱼中，每44条雄性鱼中有1条(2.2%)和22条雌性鱼中有2条(9.1%)产生GFP阳性胚胎。F1代胚胎GFP阳性率在1.4%~10.5%之间。所有F1转基因后代在孵化前均观察到母体和受精卵的表达。对F2代基因组DNA的Southern杂交分析和聚合酶链式反应分析表明，转基因已整合到寄主基因组中，并以孟德尔方式传递。这是首次通过显微注射方法在海洋鱼类的中上层产卵中建立转基因菌株。在海水养殖鱼类中建立转基因方法将是通过引入控制广泛生物现象的基因来改善其具有商业价值的特征的有力工具。
Genetic modification is an important technology for the global food production of foodstuffs. Transgenic fish strains can be established by microinjection, but this powerful method has not been developed in marine fishes due to the difficulties associated with handling their small and fragile pelagic eggs and larvae. The present paper reports the production of a transgenic strain of Nibe croaker Nibea mitsukurii (Sciaenidae), a marine fish that produces small, pelagic eggs. A microinjection technique was used to inject enhanced Green fluorescent protein (eGfp) gene driven by rainbow trout heat-shock-cognate 71 (hsc71) promoter/enhancer directly into fertilized eggs. Of 610 microinjected eggs, 498 (81.6%) survived to the 14-somite stage (24h post-fertilization) and of these, 426 (69.8%) showed highly mosaic transient expression of GFP and were selected for rearing to sexual maturity. Of the surviving fish, one out of 44 males (2.2%) and 2 out of 22 females (9.1%) produced GFP-positive embryos. Between 1.4 and 10.5% F1embryos were GFP-positive. Both maternal and zygotic expression of the eGfp gene was observed in all F1transgenic offspring prior to hatching. Southern blot analysis and PCR analysis of the genomic DNA of F2offspring revealed that transgenes were integrated into the host genome and were transmitted in a Mendelian fashion. This is the first time that a transgenic strain has been established via the microinjection method in pelagic egg spawning marine fish. The establishment of genetic modification methods in marine aquaculture fishes would be a powerful tool for improving their commercially valuable traits by introducing genes that control a wide range of biological phenomena.