成熟砂梨果实木葡聚糖转移酶基因PpXTH1的克隆及其在夏季货架期的表达规律  

Cloning of an Xyloglucan Endotransglycosylase/Hydrolase Gene(PpXTH1) from Mature Sandy Pear Fruit and Its Expression Characteristics during Summer Shelf Life

作　　者：丛郁[1] 刘洪[1] 李慧[1] 颜志梅[1] 俞明亮[1] 常有宏[1] 

机构地区：[1]江苏省农业科学院园艺研究所,江苏南京210014

出　　处：《江苏农业学报》2010年第1期143-151,共9页Jiangsu Journal of Agricultural Sciences

基　　金：江苏省博士后基金项目(070213C);江苏省农业科学院博士后基金项目(6510613);江苏省科技基础设施建设计划(BM2008008);国家农业科技成果转化基金项目(2008GB2C100100)

摘　　要：木葡聚糖内糖基转移酶（XTH）通过降解细胞壁中半纤维的主要成分木葡聚糖在果实的软化过程中发挥重要作用。为探明XTH基因与砂梨品种翠冠果实软化之间的关系，本研究以翠冠果肉cDNA为模板，采用RT．PCR和RACE方法，克隆获得了编码砂梨XTH基因的全cDNA序列（PpXTH1，1300bp）。序列分析表明，PpXTH1的5’端和3’端非翻译区分别为125bp和293bp，它的开放阅读框为882bp，编码294个氨基酸，推导的PpXTHl蛋白序列含有XTH蛋白的催化活性部位DEIDFEFL。砂梨PpXTH1与其他植物果实中的XTH蛋白具有较高的同源性，其中与苹果MdXTH1的同源性为98．0％；与猕猴桃AdXTH5的同源性为85．0％；与番茄LeXTH4的同源性为67．6％。分子进化树分析表明，PpXTHX与MdXTHl、PeXTHl、AdXTH5、PtrXTH16A同分在一组，而这些XTH蛋白与果实成熟和细胞壁的次生结构代谢密切相关。在夏季货架期，28～32℃条件下，翠冠果肉中PpXTHI的表达量持续上升，贮藏8d时其表达量最高，之后缓慢下降，1-MCP对PpXTH1的表达起延缓作用。以上结果说明本研究所克隆的基因为砂梨XTH基因家族成员，在贮藏软化过程中它的表达受乙烯调节。Xyloglucan endotransglycosylase/hydrolase （XTH） plays an important role in fruit softening through degrading xyloglucan which is the predominant hemicellulose of cell wall. To reveal the relationship between fruit softening and XTH gene during sandy pear （Pyrus pyrifolia Nakai cv. Cui Guan） fruit ripening, a full length cDNA （ 1 300 bp） of PpXTH1 was isolated from mature sandy pear fruit using RT-PCR and RACE （ rapid amplification of cDNA ends） methods. Sequence analysis showed that the isolated cDNA contained a 125 bp 5＇ untranslated region, a 293 bp 3＇ untranslated region and an 882 bp open reading frame which encoded a polypeptide including 294 amino acids. The protein induced by PpXTH1 had XTHs catalytic domain DEIDFEFL motif. The results also exhibited that PpXTH1 was 98.0%, 85.0% and 67.6% identical with the XTH protein sequence, separated from Malus demostic, Actinidia deliciosa and Lycopersicon esculentum, respectively. Molecular phylogenetic tree analysis showed that PpXTH1 with MdXTH1, PcXTH1, AdXTH5 and PtrXTH16A were divided into a group and they were all closely related to fruit ripening and cell wall secondary structure metabolism. Under high temperature shelf life condition （28 - 32 ℃ ）, expression level of PpXTH1 in sandy pear pulp rose constantly, reached the highest at 8 d, and then declined slowly. After treatment with 1- MCP, the expression level an ethylene receptor inhibitor was decreased. All these results suggest that PpXTH1 is the member of sandy pear XTH gene family and is involved in pulp softening, moreover, its expression level is regulated by ethylene during fruits softening.

关 键 词：砂梨 木葡聚糖内糖基转移酶(XTH) 基因克隆 序列分析 表达特性 

分 类 号：Q78[生物学—分子生物学]