类钙调素互作蛋白激酶基因VyCIPK1调控中国野生燕山葡萄抗旱/盐功能研究

Drought stress and soil salinization restrict the growth and development of grape, and seriously affect the yield and quality of the berries，so the research and utilization of drought/salt gene can expand the scope of the grape cultivation and improve the quality. CIPK, a kind of serine/threonine kinase, interacts with Ca ion sensors CBL to form the plant calcium signaling network, which plays a very important role in the drought, high salt, low potassium and other stresses. We first study the feature and function of CIPK genes from Chinese wild grapes Vitis. Yanshanensis which possesses high drought / salt resistance. The main contents: the localization of VyCIPK1 gene in cell、tissue and organ; Identification of kinase activity of VyCIPK1 gene; the functional study of VyCIPK1 in drought and salt resistance in non-drought resistance Arabidopsis thaliana; transformation of VyCIPK1 into Vitis vinifera Carignan and determination of function. Through above research, we determine the role in the process of signal transduction genes in drought/salt of VyCIPK1, which provides genetic resources and the basis for the function study.

干旱胁迫和土壤盐渍化制约葡萄的生长发育，严重影响浆果的产量和品质，因此研究和利用抗旱/盐基因是扩大葡萄栽培范围，提高品质的重要科学问题。CIPK是一类丝氨酸/苏氨酸激酶，与Ca离子感受器CBL相互作用形成植物钙信号传递网络，其在干旱、高盐、低钾等生物逆境的信号传递中起着极为重要的作用。本研究以高抗旱/盐的野生燕山葡萄为材料，首次研究野生燕山葡萄CIPK基因的特性与功能。主要研究内容为：研究燕山葡萄VyCIPK1基因在细胞及不同组织器官的定位；鉴定VyCIPK1基因的激酶活性；转化模式植物拟南芥，研究VyCIPK1基因的抗旱和抗盐功能；并进一步转化不抗旱的欧洲葡萄品种佳利酿，确定其功能。通过研究明确燕山葡萄VyCIPK1基因在抗旱/盐过程中的作用，为利用燕山葡萄进行抗逆育种提供基因资源，为燕山葡萄CIPK基因的调控功能的利用提供依据。

中国野生燕山葡萄具有极强的抗旱和抗盐性，但其抗逆机制不清楚。本研究从燕山葡萄中克隆得到3个类钙调素互作蛋白激酶基因，即VyCIPK1，VyCIPK2和VyCIPK3。亚细胞定位表明，VyCIPK1，VyCIPK2和VyCIPK3均定位于细胞核，属于核蛋白；逆境处理下，VyCIPK1基因主要受干旱和高盐的诱导，VyCIPK2和VyCIPK3主要受高盐的诱导；激素处理下， VyCIPK1，VyCIPK2和VyCIPK3均主要受茉莉酸甲酯（MeJA）和ABA的诱导。将VyCIPK1基因转化本氏烟草，植株产生矮化表型，通过对7种内源激素分析，导致植株矮化的主要原因可能在于生长素IBA的减少，其次SA，ABA的减少和JA的增加也有可能导致植株的矮化。种子发芽实验表明VyCIPK1基因增强了对ABA的敏感性，增强了对甘露醇的抗性；控水处理下，VyCIPK1基因可以通过调控SOD，POD，CAT等酶的活性来降低活性氧H2O2和MDA的含量，来增强对干旱的抗性。盐胁迫下，VyCIPK1基因增强了种子的萌芽率，提高了幼苗的株高，鲜重和根的数目；高盐处理下，VyCIPK1基因可以通过调控SOD，POD，CAT等酶的活性来降低活性氧H2O2、MDA的含量和电导率，增强植株的蛋白含量和叶绿素含量，从而提高了对高盐的抗性。以上表明VyCIPK1是一个优质的抗旱和抗盐基因，这位葡萄的抗逆育种提供了基因基础。