Curtis Ian S
Centre for Plant Sciences, Faculty of Biological Sciences, University of Leeds, The United Kingdom.
Methods Mol Biol. 2006;343:449-58. doi: 10.1385/1-59745-130-4:449.
Lettuce is a globally important leafy vegetable with the United States being the largest world producers. The crop is susceptible to a number of viruses that are aphid transmitted and also highly vulnerable to post harvest diseases. Although wild species of lettuce are an important source of disease resistance genes, their introgression into commercial lettuce has been limited owing to sexual incompatibilities. Hence, the development of a gene transfer system for lettuce would be extremely valuable both in improving the genetic diversity of the crop and also for the transfer of useful agronomic traits. This chapter describes an Agrobacterium-mediated gene delivery system that is highly adaptable for the production of transgenic plants using a wide range of lettuce germplasms. The system described, commonly referred to as the genotype-independent transformation system, has been used for the transfer of several agriculturally useful traits into commercial varieties of lettuce. In this case, A. tumefaciens strain LBA4404 carrying a binary vector with supervirulent pToK47 was used for infecting excised cotyledonary explants. The plant selectable marker gene neomycin phosphotransferase II (nptII) was used, and transformed plants were selected using kanamycin in the culture medium. The beta-glucuronidase gene with intron (gus-intron) was also used in the gene transfer study to confirm the transgenicity of regenerated plants further.
生菜是一种在全球范围内都很重要的叶菜类蔬菜,美国是世界上最大的生产国。这种作物易受多种由蚜虫传播的病毒影响,并且采后病害也极为脆弱。尽管野生生菜品种是抗病基因的重要来源,但由于有性不亲和性,它们向商业生菜品种中的渗入一直有限。因此,生菜基因转移系统的开发对于提高作物的遗传多样性以及转移有用的农艺性状都将极具价值。本章描述了一种农杆菌介导的基因传递系统,该系统高度适用于使用多种生菜种质生产转基因植物。所描述的系统通常被称为基因型无关转化系统,已被用于将多种农业上有用的性状导入生菜商业品种中。在这种情况下,携带带有超毒力pToK47的二元载体的根癌农杆菌菌株LBA4404被用于感染切除的子叶外植体。使用植物选择标记基因新霉素磷酸转移酶II(nptII),并在培养基中使用卡那霉素筛选转化植株。带有内含子的β-葡萄糖醛酸酶基因(gus-intron)也被用于基因转移研究,以进一步确认再生植株的转基因性。