葡萄CIPK基因家族的鉴定表达分析
Genome-wide Identification and Expression Analysis of the CIPK Gene Family in Grape

以水稻、玉米、拟南芥中已知CIPK基因注册序列为基础，从葡萄基因组数据库中电子克隆出16条CIPK基因。对其理化性质分析表明，除 VvCIPK10编码的251个氨基酸数目外，其余氨基酸数目基本稳定为300~470。整个CIPK家族的理论等电点为6~9。基因结构分析表明， VvCIPK01、 VvCIPK03、 VvCIPK04、 VvCIPK08、 VvCIPK09、 VvCIPK13包含外显子数都大于10； VvCIPK02、 VvCIPK05、 VvCIPK06、 VvCIPK07、 VvCIPK10、 VvCIPK11、 VvCIPK12、 VvCIPK14、 VvCIPK15、 VvCIPK16包含外显子数都小于7。聚类分析表明，CIPK基因被分为4个亚族，且在每一个亚族中都包含葡萄和拟南芥的CIPK基因家族成员，说明它们具有很高的同源性。对CIPK蛋白质的二级结构分析表明，葡萄CIPK基因家族所编码的蛋白质均以α-螺旋和不规则卷曲为主，而β-转角最少。亚细胞定位后发现， VvCIPK基因在细胞质中表达最多。对葡萄CIPK基因家族上游2 kb区域顺式作用元件分析表明， VvCIPK13对于ABA和脱水胁迫的响应最为明显，葡萄CIPK基因家族对于MYB转录因子和WRKY转录因子均有响应。 荧光定量分析表明， VvCIPK15在根中表达量最多，在茎中表达量最少；在不同处理下该基因表达差异性显著。其中，受PEG、ABA、NaCl诱导后呈明显上调表达，其表达量依次为PEG>NaCl>ABA。同时该基因在受到高、低温胁迫时表达量也有明显上调，9个处理中只有在山梨糖中呈下调表达。推测该基因能够参与调控干旱、盐碱、低温等逆境过程。
This work was based on known genes of rice, maize and arabidopsis. A total of 16 members of this family were identified in grape genome. Analyzing their physical and chemical properties revealed all of the genes of amino acid number distribute on 300-470 but VvCIPK10 which has 251 amino acid. Theoretical isoelectric point of the whole genes distributed on 6-9.Analyzing structure of genes indicated that a part of exon of VvCIPK were more than 10( VvCIPK01, VvCIPK03, VvCIPK04, VvCIPK08, VvCIPK09 and VvCIPK13), the others were less than 7( VvCIPK02, VvCIPK05, VvCIPK06, VvCIPK07, VvCIPK10, VvCIPK11, VvCIPK12, VvCIPK14, VvCIPK15 and VvCIPK16).Phylogenetic analysis indicated that all the proteins fell into four major clusters, and every cluster contained a part of genes of grape and arabidopsis. It indicated high homology relationship of those genes.Analyzing the secondary structure of VvCIPK protein sequence indicated that most ratio were alpha helix and irregular curly, the ratio of beta angle was the least. The whole genes chief expressed in cytoplasm by using subcellular location prediction. Analysis Cis-elements in the 2 kb upstream region indicated that the response of ABA and drought stress in VvCIPK13 was the most. Meanwhile the response of MYB and WRKY in VvCIPK were existence. Real-time quantitative analysis revealed that the expression of VvCIPK15 was the most in root. Meanwhile the expression of VvCIPK15 existed large difference under the different treatment.The inducible expression level of VvCIPK15 was significantly up-regulated under the PEG, NaCl, ABA stress. The expression was the most under the treatment of PEG. Second was NaCl. Third was ABA. The inducible expression level of VvCIPK15 was also significantly up-regulated under high or low temperature stress. This gene was only down-regulated in sorbose. VvCIPK15 may