干旱胁迫下发菜蔗糖合成酶基因Sus2克隆与差异表达  

Clone and Differential Expression of Sucrose Synthase Gene Sus2 from Nostoc flagelliforme under Drought Stress

作　　者：李晓旭[1] 丁苗苗[1] 刘阳[1] 王猛 严奉坤[2] 梁文裕[1,3] 徐婷婷[1] 王俊[4] 王玲霞[1] Li Xiaoxu;Ding Miaomiao;Liu Yang;Wang Meng;Yan Fengkun;Liang Wenyu;Xu Tingting;Wang Jun;Wang Lingxia(School of Life Sciences,Ningxia University,Yinchuan,750021;Key Lab of Ministry of Education for Protection and Utilization of Special Biologi-cal Resources in Western Yinchuan,750021;School of Agriculture,Ningxia University,Yinchuan,750021;College Education for Nationalities,Ningxia University,Yinchuan,750021)

机构地区：[1]宁夏大学生命科学学院,银川750021 [2]宁夏大学农学院,银川750021 [3]西部特色生物资源保护与利用教育部重点实验室,银川750021 [4]宁夏大学民族预科教育学院,银川750021

出　　处：《分子植物育种》2018年第20期6662-6669,共8页Molecular Plant Breeding

基　　金：国家自然科学基金项目(31660114;31660066);宁夏大学引进人才科研项目(BQD2015009)共同资助

摘　　要：蔗糖合成酶(Sus2)是调控蔗糖代谢的关键酶。为了解发菜Sus2分子信息及其对干旱胁迫的响应,设计特异性引物克隆发菜Sus2全长并进行序列分析和原核表达,分析干旱胁迫下发菜Sus2在转录水平上表达变化和蔗糖含量变化。结果表明,发菜Sus2基因全长312 bp (GeenBank登录号为MG598619),与点形念珠藻的Sus2基因及其推译氨基酸的序列相似性分别为94%和93%。推译氨基酸在第81位的Ile疏水性最强,在第27位的Gln亲水性最强。Thr有3个磷酸化位点,Ser有12个磷酸化位点,Tyr有3个磷酸化位点。Sus2蛋白二级结构及三级结构由α螺旋、β折叠、随机卷曲和β转角共同构成。将Sus2基因进行原核表达,得到一个12.29 kD的外源蛋白,LC-MS/MS分析证明该外源蛋白为蔗糖合成酶。干旱胁迫下发菜Sus2在转录水平的表达量和蔗糖含量均逐渐增加。研究结果为进一步认识发菜Sus2的分子信息以及干旱胁迫下发菜蔗糖代谢的分子机理提供了科学依据。Sucrose synthase is one of the key enzymes in regulating sucrose metabolism. In order to understand molecular information of Sus2 from Nostoc flagelliforme and its response to drought stress, specific primers were designed to clone the full length of Sus2. Also, sequence analysis and prokaryotic expression were carried out. The changes of expression and sucrose content of Sus2 on transcriptional level under drought stress were analyzed. The results showed that the full-length of Sus2 was 312 bp （GenBank access number was MG598619）. The similarity of nucleotide sequence and deduced amino acid sequence of Sus2 from N. flagelliforme and N. punctiforme PCC 73102 was 94% and 93%, respectively. In deduced amino acid, Ile was the most hydrophobic in site 81, while Gin was the most hydrophilic in site 27. Thr, Ser and Tyr had 3, 12 and 3 phosphorylation sites, respectively. Se- condary structure and tertiary structures of Sus2 protein mainly consisted of a-helix, β-sheet, β-turn and random coil. Prokaryotic expression of Sus2 gene obtained a 12.29 kD heterologous protein, which was approved to be a sucrose synthetase by LC-MS/MS analysis. Sus2 expression on the transcriptional level and sucrose content both gradually increased in N. flagell^brme under drought stress. The results provided scientific reference /br further research on molecular information of Sus2 and molecular mechanism of sucrose metabolism in N. flage lliforme under drought stress.

关 键 词：发菜 蔗糖合成酶 克隆 差异表达 干旱胁迫 

分 类 号：Q943.2[生物学—植物学]