菊花‘神马’转基因阳性株系的筛选
毕业论文关键词：菊花；CmJAZ1；叶盘法；转基因
Screening of Positive Transgenic Lines in Chrysanthemum ‘ Jinma’
Abstract: In this study, chrysanthemum cultivar ‘Jinba’ was used as the experimental material and the ORF of CmJAZ1 gene, 564bp, was cloned by high fidelity PCR. The part of ORF with Jas domain deleted, 1-348bp, was constructed into chrysanthemum expression vectors pMDC32 and the three overexpressed pMDC32-CmJAZ1△jas transgenic chrysanthemum positive lines were obtained by Agrobacterium-mediated transformation. The genomic DNA of the overexpression lines was extracted by SDS method, specific primers for Hyg resistance were selected and the plant expression vectors were successfully integrated into the plant genome identified by PCR. RNA of positive plants identified by PCR was extracted to synthesize cDNA by reverse transcription, and CmJAZ1△jas gene has been successfully expressed in transgenic lines identified by qRT-PCR.
Key words: chrysanthemum; CmJAZ1; Agrobacterium-mediated transformation; transgenic
目 录
摘要1
关键词1
Abstract…1
Key words1
引言…1
1材料与方法 2
1.1 植物材料、表达载体和菌株 …2
1.2菊花CmJAZ1基因的克隆2
1.3菊花表达载体构建 …3
1.4农杆菌叶盘法进行菊花‘神马’的遗传转化 4
1.5转基因株系的分子检测 4
1.5.1 PCR检测 …4
1.5.2 转基因株系的CmJAZ1△jas基因表达水平检测 …5
2结果与分析 7
2.1转基因菊花阳性株系的获得及分子鉴定 …7
2.1.1转基因菊花阳性株系的获得 …7
2.1.2转基因菊花的分子检测 …7
3讨论 …9致谢…10
参考文献10
菊花‘神马’转基因阳性株系的筛选
植物完整的生命周期包括营养生长和生殖生长两个阶段，其中由营养生长向生殖生长转变的过程称为开花诱导。高等植物生活史上的一个重要过程以及植物生殖发育过程中最重要的标志即开花。这一过程包括开花诱导、信号传递、属性决定、器官发生（Sun et al. 2009），涉及调控机理、不同发育方式的转换,各种不同花器官的发生和发育,以及在外界环境条件的作用下,内部信号的产生、传递和相互作用等（Yong et al. 2000）。
成花转变过程由植物自身遗传因子和外界环境因素两方面共同决定，并受错综复杂的网络信号传导途径调控（Sun et al. 2007）。近年来利用分子遗传学方法对拟南芥开花突变体进行研究表明存在四条主要的开花诱导途径：光周期途径(photoperiod pathway)、春化途径(vernalization pathway)、赤霉素途径(GA pathway)和自主途径(autonomous pathway)(Simpson and Dean, 2002; Komeda, 2004)，其中光周期途径较其他途径而言是目前研究得较多、较深入的一条途径。随着分子生物学及分子遗传学的迅猛发展，许多与光周期途径开花相关的基因已经被发现并克隆。