摘要
为了解腺苷二磷酸葡萄糖焦磷酸化酶(AGPase)基因的功能及其与油菜种子油脂含量变化之间的关系,通过构建2类特异启动子-基因的表达载体:pMB-DOF-Glgc(含拟南芥球形胚阶段启动子DOF及大肠杆菌AGPase突变基因Glgc 16)和pMB-Cruc-Glgc(含油菜种子特异蛋白启动子Cruc和Glgc 16),获得转基因甘蓝型油菜株系,分别在胚胎早、晚期特异过表达外源AGPase基因Glgc,拟增加油菜胚淀粉的合成,促进脂肪酸的合成,达到增加油菜种子含油量的目的。利用qPCR方法,从两组转化植株中筛选出10个单拷贝阳性株,分析不同发育时期种子淀粉和含油量。结果表明,DOF:Glgc和CruC:Glgc转化体种子淀粉含量显著增加,且增加的峰值与启动子时空特异性一致。具体表现为:从淀粉含量变化看,DOF:Glgc组和CruC:Glgc组种子平均相对淀粉含量分别从发育前期(22 DAF,开花后22 d)和发育中期(35 DAF)开始较对照增加;而对照淀粉积累峰值均出现在31 DAF,分别是9.43%和9.76%;DOF:Glgc组的峰值也出现在31 DAF,达12.33%;CruC:Glgc组出现在35 DAF,达12.92%,相反地,两组对照成熟种子平均含油量分别为40.12%和40.00%,两组转基因组则为40.35%和40.26%。测定已获得的150株转基因阳性植株的种子含油量,结果显示:DOF:Glgc组和CruC:Glgc组平均含油量分别为40.58%和40.60%,空载体转基因对照的受体材料平均含油量分别为40.30%和40.33%,种子含油量增加并不显著。综上结果提示,增加油菜胚中的淀粉含量可能无法改变最终碳流的分配从而影响种子含油量。
In order to understand the function of adenosine diphosphate glucose pyrophosphorylase(AGPase)gene and its relationship with the changes in oil content of rapeseeds,two types of specific promoter-gene expression vectors were constructed:pMB-DOF-Glgc(overexpress the exogenous AGPase gene Glgc in early embryos)and pMB-Cruc-Glgc(overexpress the exogenous Glgc in late embryos).By detection of transgenic positive plants subsequently,it was intended to increase starch synthesis in embryo,promote synthesis of fatty acids,and achieve the purpose of increasing the oil content of rape seeds.Ten single-copy positive strains were selected from the transformed positive plants in each vector group by qPCR method.The relative starch content in seeds at different developmental stages and oil content of mature seeds of the above positive plants were analyzed.Oil content analysis was performed on mature seeds of all transformed positive plants.Results presented that the average relative content of the starch in single-copy plant seeds in DOF:Glgc group and Cruc:Glgc group increased from the pre-development stage(22 DAF)and the middle stage development(35 DAF),respectively.The peaks of starch accumulation in two mock-vehicle controls were 9.43%and 9.76%,at 31 DAF respectively.Maximum starch content of the DOF:Glgc group also appeared at 31 DAF,reaching 12.33%.While in Cruc:Glgc group it appeared at 35 DAF,reaching 12.92%.The average oil content of mature seeds in the two control groups was 40.12%and 40.00%,and the average oil content of mature seeds in the DOF:Glgc group and Cruc:Glgc group were 40.35%and 40.26%,respectively.For 150 transgenic lines,oil content of two Cruc:Glgc group were 40.58%and 40.60%respectively,while the average oil content of the controls were 40.30%and 40.33%.The analysis results revealed that the starch content of DOF:Glgc and Cruc:Glgc transformants significantly increased,and the peak value of the increase was consistent with the promoter′s spatiotemporal specificity.However,the increase in oil content
作者
赵凯琴
杨清辉
张云云
罗延青
王敬乔
李劲峰
陈苇
田正书
ZHAO Kai-qin;YANG Qing-hui;ZHANG Yun-yun;LUO Yan-qing;WANG Jing-qiao;LI Jing-feng;CHEN Wei;TIAN Zheng-shu(College of Agronomy and Biotechnology,Yunnan Agricultural University,Kunming 650201,China;Industrial Crops Research Institute,Yunnan Academy of Agricultural Sciences,Kunming 650205,China)
出处
《中国油料作物学报》
CAS
CSCD
北大核心
2021年第3期426-434,共9页
Chinese Journal of Oil Crop Sciences
基金
国家重点研发计划项目(2016YFD0101305)。
关键词
甘蓝型油菜
腺苷二磷酸葡萄糖焦磷酸化酶
转基因
淀粉
含油量
Brassica napus L.
adenosine diphosphate glucose pyrophosphorylase(AGPase)
transgene
starch
oil content
