园艺作物育种学、第九章、生物技术育种2.ppt

第九章 生物技术育种 （一）目的基因的获取 　 基因组DNA文库（genomic DNA library） 指将某生物的全部基因组DNA切割成一定长度的DNA片断，克隆到某种载体上而形成的集合。 （二）植物表达载体的构建 一些常见的遗传选择标记和报告基因（Reporter Genes/Selectable Markers） （三）植物遗传转化方法 1、农杆菌介导的遗传转化 2、基因枪介导的遗传转化 3、花粉管通道法 Agrobacterium tumefaciens mediated plant transformation （四）转化细胞的筛选及转基因植株鉴定 （五）外源基因的整合及表达分析 通常外源基因在受体植物染色体上是随机插入。也有一些报告表明有时特定的外源基因插入位点在序列上有一定的倾向性，并不完全随机。外源基因插入植物染色体的拷贝数差异较大，插入过程中可能会发生结构变化。 外源基因在转化体中的表达可以通过Western杂交和ELISA分析来检测。 （六）转基因作物新品种的培育 作物转基因育种可以通过转基因方法直接培育出优良新品种，包括外源目的基因直接导入优良推广品种或品系以改良个别性状，或者是将转基因作物作为亲本材料在作物品种培育过程中间接利用。 植物基因工程育种与常规育种是相辅相成的，基因工程必须与常规育种密切配合，通过杂交或回交育种手段，经大田试验，从而加快优良新品种（系）的培育进程。 二、转基因技术在育种上的应用 1. New horticultural varieties Understand the anthocyanin (pigment) biosynthetic pathways, can produce novel flower varieties 2. Improve pest resistance Transfer gene that inhibits digestion of starch (amylase inhibitor) from bean to garden pea Stops attack by weevils Transfer one of many Cry genes from Bacillus thuringiensis to plants Makes protein (Bt) toxins, only small amount needed in plant to kill insect pests 2. Improve pest resistance (cont’d) Transfer mutated gene in shikimic acid pathway from E. coli to make resistant (Roundup Ready) plants Glyphosate herbicides inhibit an important enzyme in this pathway; plants need aromatic AAs to grow! 3. Improve nutritive value of plants Use metabolic engineering to insert new pathways into plants or improve expression of enzymes in existing ones.Most crop plants are deficient in one or more amino acids maize is low in lysine, methionine and tryptophan Improve vitamin quality of crops “Golden rice” higher in beta-carotene, the precursor to vitamin A Improve value of feed crops Transfer a fungal enzyme (phytase) to crops to remove phytic acid from feed and improve phosphate availability 4. Improve resistance to stress e.g., salt stress: Express high levels of Na+/H+ antiporter in vacuole membrane This allows plants to grow in high Na (50X normal limit)