枳与克里曼丁橘落叶相关生理变化及候选基因功能分析

发布时间：2018-09-11 18:21

【摘要】：柑橘作为世界第一大果树,是典型的亚热带植物。在柑橘类植物中只有枳属(Poncirus trifoliata)具有落叶性。落叶性在植物的生长发育过程中起着重要的作用,如促进休眠,减少蒸腾作用,提高抗寒性等。近年来,有文章以常绿柑橘为材料研究干旱或乙烯处理等逆境条件下的离层形成与叶片脱落现象,而以枳属为试材研究落叶机理的文章少之又少。因此,弄清控制柑橘落叶的原因及其内在基因调控机理具有重要的意义。本试验以早实枳和克里曼丁橘为主要研究材料,探究枳不同于常绿柑橘的一种与季节变换相适应的主动性落叶抗寒机制。主要研究结果如下:1.利用石蜡切片技术对早实枳和克里曼丁橘的一年生茎和叶进行横切,观察茎和叶的维管束(导管)结构差异,发现早实枳的导管管径、导管数量以及宽导管比例均比克里曼丁橘小,说明早实枳的导管吸水能力比克里曼丁橘弱,但其窄导管输导水分更安全,更有利于适应低温逆境环境抵御冬季严寒;2.对早实枳和克里曼丁橘的一年生枝条在落叶前后进行树体生理指标的测定,即相对电导率变化、丙二醛含量、脯氨酸含量、可溶性糖含量以及可溶性蛋白含量的测定,以观察两品种在落叶前后的树体生理指标变化和抗寒力变化。结果表明在自然温度变化下,早实枳比克里曼丁橘更能适应低温环境;3.本实验室前期通过QTL定位、基因功能注释以及对枳幼苗低温处理进行qRT-PCR共筛选出了30个落叶候选基因,从中挑选出5个基因构建超表达载体,并转到模式植物番茄中异位表达,以进行该基因的功能验证。结果表明超表达转基因株系35S::ABS3和35S::ABS7比野生型对照植株更能适应低温环境。说明ABS3和ABS7基因可以响应低温诱导。4.本实验通过了解枳落叶性状的遗传规律和落叶关联基因的功能,以进一步认识落叶植物不同于常绿植物的主动适应气候变换,获得在寒冷地区生存能力的特性,为进一步研究柑橘抗寒分子育种提供重要的理论基础。
[Abstract]:Citrus, as the largest fruit tree in the world, is a typical subtropical plant. Among citrus plants, only (Poncirus trifoliata) has deciduous properties. Deciduous property plays an important role in plant growth and development, such as promoting dormancy, reducing transpiration and improving cold resistance. In recent years, evergreen citrus has been used to study the formation of layer separation and leaf shedding under drought or ethylene treatment, but there are few articles on the deciduous mechanism of trifoliate (Trifoliate orange). Therefore, it is of great significance to understand the cause of controlling citrus leaves and its intrinsic gene regulation mechanism. In this study, we studied the active deciduous cold resistance mechanism of trifoliate orange, which is different from evergreen citrus, and adapted to seasonal change. The main results are as follows: 1. The annual stems and leaves of precocious trifoliate orange and Cleimanthus trifoliate were crosscut by paraffin section technique. The differences of vascular bundle (vessel) between stem and leaf were observed and the diameter of ductal tube of precocious trifoliate orange was found. The number of ducts and the ratio of wide ducts were smaller than those of Creman's, which indicated that the ducts of precocious trifoliate orange were weaker in water absorption than those of Cleimanthus, but its narrow conduits were safer to transport water, which was more advantageous to adapt to the low temperature environment and resist the cold in winter. The physiological indexes of the annual branches of precocious trifoliate and Crymannia were measured before and after the fallen leaves, namely the relative electrical conductivity, the content of malondialdehyde, the content of proline, the content of soluble sugar and the content of soluble protein. The changes of physiological indexes and cold resistance of the two varieties before and after falling leaves were observed. The results showed that the precocious trifoliate orange could adapt to low temperature environment better than Creman's under natural temperature. A total of 30 deciduous candidate genes were screened by QTL mapping, gene function annotation and qRT-PCR treatment of trifoliate seedlings in the early stage of the experiment, and 5 genes were selected to construct superexpression vector and transferred to heterotopic expression in model plant tomato. To verify the function of the gene. The results showed that the overexpression transgenic lines 35S::ABS3 and 35S::ABS7 could adapt to low temperature environment better than wild-type control plants. These results suggest that ABS3 and ABS7 genes can respond to hypothermia induction. 4. In order to further understand the active adaptation of deciduous plants from evergreen plants to climate change and to obtain the characteristics of survival ability in cold regions, the genetic rules of defoliation traits and the functions of deciduous associated genes in trifoliate orange (Trifoliate trifoliate) were studied in this experiment. It provides an important theoretical basis for further study on cold resistant molecular breeding of citrus.
【学位授予单位】：华中农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S666

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