【作者】 汤浩茹；

【导师】 任正隆；

【作者基本信息】 四川农业大学 ， 作物遗传育种， 2000， 博士

【摘要】 核桃( Juglans regia L.)是一种适应性强、分布广、多用途的优良经济栽培树种。发展核桃生产,品种是关键。由于核桃的生物学特性的限制,用常规育种手段改良核桃品种的周期长、效率低,而用现代生物技术,通过目的性非常强的基因操作改良核桃品种不仅可以加速新品种的选育,而且还能避免因有性重组而带来的不良后果。能否获得转基因植株,除了基因的转移方法外,在很大程度上取决于有效的离体培养再生系统。叶园片培养与原生质体培养是目前应用最为广泛而又最成功的植物转基因研究的离体培养再生系统。尽管核桃的离体培养技术得到了广泛深入的研究,但是,核桃的原生质体培养与叶园片培养还未能成功。核桃的体细胞胚具有完整的两级结构,能一次再生完整的植株。同时,核桃的体细胞胚能反复产生次生胚,而且次生胚是表皮单细胞起源的。因此,核桃的体细胞胚发生为核桃的转基因研究提供了良好的离体培养再生系统。自1985年Tulecke和McGranahan首次报道核桃的体细胞胚胎发生以来,相继在核桃的五个种、三个杂种中有了体细胞胚胎发生的报道。由于核桃的体细胞胚胎发生均是在不同条件下获得的,而且真正用于核桃转基因研究的反复次生胚发生以及体细胞胚的萌发与成苗还未能得到深入研究,致使核桃转基因研究的成功报道甚少。针对这些问题,本研究采用德国主栽优良核桃品种‘Weinsberg 1’、‘No.120’以及‘No.1247’(Juglans regia L.)不同发育时期的胚珠、胚轴与子叶以及试管苗叶片、叶柄和茎段为试材系统研究了核桃的体细胞胚胎发生,首次获得了核桃胚珠、胚轴与试管苗叶柄的体细胞胚。优化了核桃的反复次生胚发生、体细胞胚的成熟、萌发与成苗,首次将脱水处理引入处理核桃,从而极显著地提高了核桃体细胞胚的萌发率和成苗率。同时探讨了以核桃体细胞胚为受体进行外源基因的转化方法,首次系更多还原

【Abstract】 Walnut, which produces both nuts and timbers, is one of the most important nut crops world-wide. Breeding of walnut by conventional methods is difficult due to its highly heterozygous characters and long generation cycles. Genetic engineering methods provide an attractive support to complement conventional breeding of walnut. However, the lacking availability of transformation protocols is still the major bottleneck in this approach. Somatic embryogenesis has been considered as the most promising method in genetic improvement not only because it is an efficient plant regeneration system but also it can make the superior genotype selection more efficiency when combined with genetic engineering and conventional methods. In walnut, somatic embryogenesis is highly important because somatic embryos are the only explant capable of regeneration after transformation.Since the first reports of somatic embryogenesis in walnut in 1985 and regeneration of transgenic walnut plants from somatic embryos in 1988, successful somatic embryogenesis and regeneration of transformed somatic embryos have been achieved. However, somatic embryogenesis and genetic transformation in walnut are still far from routine and many factors affect the frequency of somatic embryogenesis and genetic transformation. In an attempt to establish a protocol for walnut transformation, we obtained somatic embryos from various explants of English walnut (Juglans regia L.) and repetitively somatic embryo lines. By using the repetitively somatic embryos as target tissues we successed in recovery of transgenic plants after Agrobacterium tumefaciens mediation.To develop somatic embryogenesis in walnut, young intact ovules of‘No.更多还原