Council of Scientific and Industrial Research-National Botanical Research Institute, Rana Pratap Marg, Lucknow, Uttar Pradesh, India.
PLoS One. 2013 Apr 30;8(4):e63064. doi: 10.1371/journal.pone.0063064. Print 2013.
Sterol glycosyltrnasferases (SGT) are enzymes that glycosylate sterols which play important role in plant adaptation to stress and are medicinally important in plants like Withania somnifera. The present study aims to find the role of WsSGTL1 which is a sterol glycosyltransferase from W. somnifera, in plant's adaptation to abiotic stress.
The WsSGTL1 gene was transformed in Arabidopsis thaliana through Agrobacterium mediated transformation, using the binary vector pBI121, by floral dip method. The phenotypic and physiological parameters like germination, root length, shoot weight, relative electrolyte conductivity, MDA content, SOD levels, relative electrolyte leakage and chlorophyll measurements were compared between transgenic and wild type Arabidopsis plants under different abiotic stresses--salt, heat and cold. Biochemical analysis was done by HPLC-TLC and radiolabelled enzyme assay. The promoter of the WsSGTL1 gene was cloned by using Genome Walker kit (Clontech, USA) and the 3D structures were predicted by using Discovery Studio Ver. 2.5.
The WsSGTL1 transgenic plants were confirmed to be single copy by Southern and homozygous by segregation analysis. As compared to WT, the transgenic plants showed better germination, salt tolerance, heat and cold tolerance. The level of the transgene WsSGTL1 was elevated in heat, cold and salt stress along with other marker genes such as HSP70, HSP90, RD29, SOS3 and LEA4-5. Biochemical analysis showed the formation of sterol glycosides and increase in enzyme activity. When the promoter of WsSGTL1 gene was cloned from W. somnifera and sequenced, it contained stress responsive elements. Bioinformatics analysis of the 3D structure of the WsSGTL1 protein showed functional similarity with sterol glycosyltransferase AtSGT of A. thaliana.
Transformation of WsSGTL1 gene in A. thaliana conferred abiotic stress tolerance. The promoter of the gene in W.somnifera was found to have stress responsive elements. The 3D structure showed functional similarity with sterol glycosyltransferases.
甾醇糖苷转移酶（SGT）是糖基化甾醇的酶，在植物适应应激中起着重要作用，在药用植物如睡茄中具有重要意义。本研究旨在寻找睡茄甾醇糖苷转移酶（WsSGTL1）在植物适应非生物胁迫中的作用。
通过农杆菌介导的转化，使用二元载体 pBI121，通过花浸法将 WsSGTL1 基因转化到拟南芥中。在不同的非生物胁迫下，如盐、热和冷胁迫下，比较转基因和野生型拟南芥植株的表型和生理参数，如发芽率、根长、茎重、相对电解质导率、MDA 含量、SOD 水平、相对电解质泄漏和叶绿素测量。通过 HPLC-TLC 和放射性酶测定进行生化分析。使用基因组步行试剂盒（Clontech，美国）克隆 WsSGTL1 基因的启动子，并使用 Discovery Studio Ver. 2.5 预测 3D 结构。
Southern 杂交和分离分析证实 WsSGTL1 转基因植株为单拷贝。与 WT 相比，转基因植株表现出更好的发芽率、耐盐性、耐热性和耐寒性。在热、冷和盐胁迫下，转基因植物的 WsSGTL1 水平升高，同时还检测到其他标记基因如 HSP70、HSP90、RD29、SOS3 和 LEA4-5 的表达。生化分析表明形成了甾醇糖苷，并增加了酶活性。从睡茄中克隆 WsSGTL1 基因的启动子并测序后，发现它含有应激响应元件。WsSGTL1 蛋白的 3D 结构的生物信息学分析显示与拟南芥的甾醇糖苷转移酶 AtSGT 具有功能相似性。
在拟南芥中转化 WsSGTL1 基因赋予了非生物胁迫耐受性。在睡茄中，基因的启动子被发现含有应激响应元件。3D 结构显示与甾醇糖苷转移酶具有功能相似性。
