马铃薯离体块繁的实践与应用.ppt

第五章 植物离体快繁的实践与应用,植物器官培养（Organ culture ）,是指植物离体的某一器官的全部或部分或器官原基的培养，包括茎段、茎尖、球茎、叶片、子叶、花序、花瓣、子房、花药、花托、果实、种子等。,园艺植物离体繁殖,龟背竹试管苗,月季试管苗开花,非洲紫罗兰壮苗生根,月季培养腋芽萌发,马铃薯脱毒及快速繁殖,Stock plant selection and preparation,0,stage,马铃薯是一种全球性的重要经济作物。适应性广，营养价值高，耐贮藏适运输，既是一种重要的粮食作物也是一种调节市场供应的重要蔬菜。马铃薯原产于拉丁美洲，当被发现可以食用后，很快传到世界各地，成为栽培很广的一种作物。,1、形态特性 根 根系由出生根和匍匐根组成。 茎 分地上部分和地下部分。 叶 全缘，心脏形。 花 为聚伞花序。 果实 浆果。,马铃薯生物学特性,马铃薯性喜冷气候，不耐高温和霜冻。 发芽适温为1218。 地上茎叶生长温度为1721。 块茎形成要求昼温1424，夜温1214。 结薯期要求12h左右的短日照和疏松、湿润、肥沃以及通气良好的土壤条件。 主要以块茎繁殖。,2、 生物学习性 和繁殖方式,危害有17种之多，如X病毒、S病毒、Y病毒、M病毒、A病毒、花叶病毒、纺锤形块茎病毒等。 马铃薯病毒可使块茎产量减少50%80%。,马铃薯病毒的种类,Explant Genetically superior Clean environment Young and vigorous,Explant selection,Select a suitable potato mother plants (high productivity, high quality, free from diseases and so on) Grow the selected mother plants under more hygienic conditions,热处理脱毒,脱毒：室内发芽，热处理(38) 2 周；,原理： 当植物组织处于高于正常温度的环境（35-40）时，组织内部的病毒部分或全部钝化，但寄主植物的组织很少或不会受到伤害。,Treatment of virus free,Excise a shoot tips or buds from potato tubes after treatment of high temperature (38, 2 weeks) for virus-free,茎尖培养脱毒原理,1.染病植株体内病毒的分布不均匀，越靠近茎顶的感染越轻，生长点（约0.10.5）几乎不含或含病毒很少。原因是无维管束，病毒只能通过胞间连丝传递，不及细胞不断分裂和生长的速度。因此可利用微茎尖培养脱毒种苗。 2.茎尖大小与脱毒效果成反比。,Initiation of culture,1,stage,Purpose Establishment of aseptic cultures in vitro Operation Take explants from mother plants such as buds leaf pieces, flowers into the vessels.,Explant,Put on medium,Take bud under aseptic conditions,Establishment of aseptic cultures in vitro ? Medium ? Culture environment conditions?,Discussions,1. Wash the potato shoot tips for 10 min. 2. Soak them in 70% ethanol solution for 30 sec. 3. Soak them in 5% NaClO or 0.1% HgCl solution for 15 or 10 min. for surface sterilization. 4. Rinse them at least 3-5 times with sterile water,6. Excise a meristem (0.1-0.3mm) from a shoot tip 7. Place it aseptically on sterilized medium,MS+BA0.2mg +3%蔗糖 温度：2125 光照度：2000-3000 lx 光照时间：12h/d 2-3周长愈伤，4-5周长绿点，3-6月长成2-3厘米小芽，越慢越好，出芽很快的扔掉。,马铃薯初代培养：,病毒植物的鉴定,指示植物法 电子显微镜检查法 酶联免疫鉴定法,Multiplication,2,stage,马铃薯脱毒苗组培扩繁,Medium ? Culture environment conditions?,Discussions,MS+0.2BA + 10mg D-泛酸钙 +3%蔗糖； MS + 3% sugar + 0.01% C; 20-25，3000-4000LX，14-16hr光照； 每3周继代一次，单芽茎段约1厘米； 原则试管苗用2年3年。,马铃薯继代培养基：,After culturing for 3-4 weeks, elongated shoots are transferred to multiplication medium,Stage,2,6,Principle of multiplication Nodal proliferation， Axillary bud formation One nodal cutting one plant，high genetics，simple operation, higher survival ex-vitro，but lower multiplication rate at beginning.,3,6 shoots,6 shoots,Nodal cuttings,CUT,CUT,Axillary bud formation,Restrain top-bud growth, enhance formations of axilary buds High genetics，simple operation,Axillary branching,Next explant,CUT,Principle of multiplication Young shoots or bud grown on a appropriate culture medium become cluster shoots. Higher multiplication rate with higher mutation when high phytohormone used.,Shoot proliferation,Elongation and rooting,3,stage,Shoot elongation,Rooting,Elongation medium,Rooting medium,Stage 3,A. Shoot elongation B. Root induction & development,Medium ? Culture environment conditions?,Discussions,MS+NAA0.2+D-泛酸钙10mg +3%蔗糖； MS (surge-free medium); 20-25，3000-4000LX，14-16hr光照； 每3周继代一次，单芽茎段约1厘米； 原则试管苗用2年3年。,马铃薯生根培养：,伸長,At this stage, rooted plantlets are transferred to a cell tray and grown under 60% shaded and high RH conditions for 7 to 10 days to acclimatize Acclimatization are usually attained in 2 to 3 weeks,Acclimatization,4,stage,From in vitro environment to ex vitro environment From photomixotrophy to photoautotrophy,Carbon source: sugar in the media Aseptic environment Low light intensity High relative humidity,Carbon source: CO2 in the air Nonaseptic environment high light intensity Low relative humidity,In vitro environment,Ex vivo environment,Acclimatization,炼苗室温度：白天2327，夜间不低于14。 炼苗具体方法：移植前7d左右，将长有35片叶、高23