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超表达杨树SBPase基因促进拟南芥光合作用及营养生长

王俊秀 周扬颜 韩潇 安轶 郭惠红 夏新莉 尹伟伦 刘超

王俊秀, 周扬颜, 韩潇, 安轶, 郭惠红, 夏新莉, 尹伟伦, 刘超. 超表达杨树SBPase基因促进拟南芥光合作用及营养生长[J]. 北京林业大学学报, 2018, 40(3): 26-33. doi: 10.13332/j.1000-1522.20170436
引用本文: 王俊秀, 周扬颜, 韩潇, 安轶, 郭惠红, 夏新莉, 尹伟伦, 刘超. 超表达杨树SBPase基因促进拟南芥光合作用及营养生长[J]. 北京林业大学学报, 2018, 40(3): 26-33. doi: 10.13332/j.1000-1522.20170436
Wang Junxiu, Zhou Yangyan, Han Xiao, An Yi, Guo Huihong, Xia Xinli, Yin Weilun, Liu Chao. Overexpression of Populus SBPase gene promoting photosynthesis and vegetative growth in Arabidopsis thaliana[J]. Journal of Beijing Forestry University, 2018, 40(3): 26-33. doi: 10.13332/j.1000-1522.20170436
Citation: Wang Junxiu, Zhou Yangyan, Han Xiao, An Yi, Guo Huihong, Xia Xinli, Yin Weilun, Liu Chao. Overexpression of Populus SBPase gene promoting photosynthesis and vegetative growth in Arabidopsis thaliana[J]. Journal of Beijing Forestry University, 2018, 40(3): 26-33. doi: 10.13332/j.1000-1522.20170436

超表达杨树SBPase基因促进拟南芥光合作用及营养生长

doi: 10.13332/j.1000-1522.20170436
基金项目: 

国家自然科学基金项目 31570308

国家自然科学基金项目 31600484

国家自然科学基金项目 31770649

国家重点研发计划课题 2016YFD0600403

详细信息
    作者简介:

    王俊秀。主要研究方向:植物抗逆分子生物学。Email: junxiu2016@bjfu.edu.cn 地址:100083北京市清华东路35号北京林业大学林木育种国家工程实验室

    责任作者:

    刘超,助理研究员。主要研究方向:植物抗逆生理生态学。Email: liuchao1306@163.com 地址:同上

  • 中图分类号: S718.43

Overexpression of Populus SBPase gene promoting photosynthesis and vegetative growth in Arabidopsis thaliana

  • 摘要: 目的卡尔文循环是植物光合作用中极为重要的生理过程,对植物的生长发育具有显著影响。前期研究表明,高光合速率的速生欧美杨的景天庚酮糖-1, 7-二磷酸酯酶(SBPase)基因表达水平在速生期显著上调,预示该基因在光合碳固定过程中可能起着关键作用。方法为进一步解析SBPase在木本植物光合速率和生长发育中的作用,本文从速生欧美杨品系NE19中克隆得到了PdSBPase基因,并构建35S:PdSBP:GFP表达载体,采用农杆菌花序侵染法转化拟南芥,通过抗生素筛选,PCR鉴定和组织定位等多种方式鉴定并成功得到了超表达PdSBPase拟南芥株系。结果在正常生长状态下,超表达植株的叶面积、根长、株高都优于野生型和突变体,其中叶面积是野生型的1.79倍,根长是野生型的1.93倍,而突变体表现为植株矮化,叶子明显发黄短小,叶绿素含量低于野生型株系。转基因株系SBPase酶活是野生型1.4倍,是突变体的1.9倍,RuBP产量以及淀粉含量均要高于野生型和突变体株系,RuBP产量分别是野生型和突变体的1.37和1.76倍,转基因株系的淀粉含量达到了50.26μg/g,而突变体的淀粉含量未检出。结论这些结果说明,PdSBPase对RuBP的形成和淀粉等多糖的合成起到关键作用,能促进植物积累更多的碳水化合物,进而正向调控植物的光合能力。

     

  • 图  1  PdSBP与其他物种SBP的分子进化树及蛋白序列比对、35S:PdSBP:GFP植物表达载体结构图及超表达拟南芥植株PCR检测、PdSBP在NE19各器官的表达量分析

    PdSBP与其他物种SBP蛋白的分子进化树,红点标记的为本实验研究对象PdSBP;B. 35S :PdSBP :GFP植物表达载体;C.黑杨与毛果杨(PT08G06380和PT10G19330)、拟南芥SBP蛋白序列比对结果,蓝色表示相同的碱基;D.PdSBP在NE19各器官的表达量分析;E.超表达拟南芥植株PCR检测(M: marker; TG: transgenic gene; NG:negative control; PG:positive control)。

    Figure  1.  Molecular phylogenetic tree and sequence alignment of of PdSBP and other SBP proteins, 35S: PdSBP: GFP structure diagram of plant expression vector and PCR detection of overexpression Arabidopsis thaliana, expression of PdSBP in various organs of NE19

    A, molecular phylogenetic tree of PdSBP and other SBP proteins, the red dot marker is the experimental subject PdSBP; B, 35S:PdSBP :GFP plant expression vector; C, sequence alignment of SBP protein between Populus nigra and Populus trichocarpa and Arabidopsis thaliana. The same base was expressed in blue; D, expression of PdSBP in various organs of NE19; E, PCR detection of overexpression Arabidopsis thaliana.

    图  2  超表达拟南芥植株叶片GFP检测及亚细胞定位

    Figure  2.  GFP detection and subcellular localization of overexpressing PdSBP in Arabidopsis thaliana leaf cells

    图  3  4个株系叶面积和根长比较

    A、C.4个株系的根长比较,野生型Col-0、超表达型oxPdSBP#10和oxPdSBP#8、突变体sbp,下同;B、D.4个株系叶面积统计。图中数据为平均值±标准误(n=3)。4个株系间不同字母表示在P<0.05水平上差异显著。

    Figure  3.  omparison of leaf area and root length of the four strains

    A, C, root length comparison of four lines, wild type (Col-0), overexpression type (oxPdSBP#10and oxPdSBP#8), mutant(sbp), and the same below; B, D, leaf area statistics of 4 lines.Data in the figure are mean ±SE (n=3).Different letters in the four strains mean significant difference at P<0.05 level.

    图  4  4个株系茎伸长率统计

    A、B 4个株系在21~31d生长期间茎伸长率统计。图中数据为平均值±标准误(n=3)。4个株系间不同字母表示在P<0.05水平上差异显著。

    Figure  4.  Statistics of stem elongation of the four lines

    A, B, statistics in stem elongation of 4 lines during 21-31 days of growing season.Data in the figure are mean ± SE (n=3).Different letters in the four strains mean significant difference at P<0.05 level.

    图  5  正常生长条件下,4个株系植物生理指标分析

    A.4个株系在正常生长条件下的净光合速率;B.叶绿素荧光参数Fv/Fm;C.4个株系总叶绿素含量;D.叶绿素a/b的比值。图中数据为平均值±标准误(n=3)。4个株系间不同字母表示在P<0.05水平上差异显著。

    Figure  5.  Physiological analysis of four lines under normal growth conditions

    A, net photosynthetic rate of 4 lines under normal growth conditions determined; B, chlorophyll fluorescence parameters Fv/Fm; C, the ratio of total chlorophyll content; D, chlorophyll a/b in 4 lines. Data in the figure are mean ± SE (n=3). Different letters in the four strains mean significant difference at P<0.05 level.

    图  6  SBP酶活、RuBP和淀粉含量的比较

    A.4个株系在正常生长条件下RuBP含量统计;B.正常生长条件下SBPase酶活性统计;C.正常生长条件下淀粉含量统计。图中数据为平均值±标准误(n=4)。4个株系间不同字母表示在P<0.05水平上差异显著。

    Figure  6.  Comparison of SBPase activity, RuBP and starch content

    A, RuBP content of 4 lines under normal growth conditions; B, SBPase enzyme activity statistics under normal growth conditions; C, statistics of starch content under normal growth conditions.Data in the figure are mean ± SE (n=4). Different letters in the four strains mean significant difference at P<0.05 level.

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  • 收稿日期:  2017-12-07
  • 修回日期:  2018-01-02
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