茉莉花原生质体瞬时表达体系的建立及应用  

Development of a protoplast-based transient expression system and application in Jasminum sambac

作　　者：张娅 刘晓烽 张婧 何弦 袁媛[2] 吕美玲 黄晨星[1] 郑祥梓[1] 缪颖[1] 伍炳华[2] ZHANG Ya;LIU Xiaofeng;ZHANG Jing;HE Xian;YUAN Yuan;LV Meiling;HUANG Chenxing;ZHENG Xiangzi;MIAO Ying;WU Binghua(Fujian Provincial Key Laboratory of Plant Functional Biology,College of Life Sciences,Fujian Agriculture and Forestry University;College of Horticulture,Fujian Agriculture and Forestry University,Fuzhou,Fujian 350002,China)

机构地区：[1]福建农林大学生命科学学院,福建省植物功能生物学与绿色农业重点实验室 [2]福建农林大学园艺学院,福建福州350002

出　　处：《福建农林大学学报（自然科学版）》2019年第6期727-735,共9页Journal of Fujian Agriculture and Forestry University:Natural Science Edition

基　　金：福州市校(院所)科技合作项目(2017-G-84);园艺学院国家级创新训练项目(201810389035);福建省自然科学基金(2015N0019);国家自然科学基金(31701862);福建农林大学“校杰出青年科研人才”计划项目(xjq201717)

摘　　要：探索了茉莉花原生质体瞬时转化的方法,发现室外种植盛开的茉莉花花瓣更适于分离有活力的原生质体,在15 g·L^-1纤维素酶、8 g·L^-1果胶酶和4 g·L^-1离析酶共同作用下,于25℃黑暗条件酶解5 h,所获得的原生质体产量和活性最佳.当细胞浓度为4×10^5个·mL^-1,外源质粒DNA含量为75μg·mL^-1原生质体时,在200 g·L^-1的PEG-4000介导转化2~10 min,可获得较高的转化效率.利用本研究建立的茉莉花原生质体瞬时转化体系成功鉴定了3个茉莉花来源蛋白的亚细胞定位,为高通量开展茉莉花关键基因的功能研究提供了技术支持.A method was proposed to transiently express proteins in Jasminum sambac protoplasts.The results showed that petals from full-bloomed flowers grown under natural condition are more favorable for protoplast isolation.The optimal isolation condition is to digest petals in the enzyme solution containing 15 g·L^-1 cellulase,8 g·L^-1 pectinase and 4 g·L^-1 macerozyme for 5 h at 24℃in the dark,yielding petal protoplasts with satisfactory yield and vitality.High transformation efficiency of around 70%can be achieved when the protoplasts in a concentration of 4×10^5 cells per milliliter solution mixed with exogenous plasmid DNA in a ratio of 75μg·mL^-1 protoplasts,followed by the mediation of 200 g·L^-1 PEG-4000 for 2-10 min.Furthermore,subcellular localizations of three proteins derived from jasmine were successfully identified in jasmine protoplasts.This protoplast-based transient expression system provides technical support for the rapid functional characterization of key genes in J.sambac.

关 键 词：茉莉花 原生质体 亚细胞定位 

分 类 号：S685.16[农业科学—观赏园艺]