<graphic xmlns:xlink="http://www.w3.org/1999/xlink" xlink:href="0496-3490-41-3-507/img_1.png"/>稗草(<em> Echinochloa crusgalli</em>)根型<em> ppc</em>基因对水稻的遗传转化及其对光合速率的调节效应

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张桂芳, 丁在松, 赵明. 稗草( Echinochloa crusgalli)根型 ppc基因对水稻的遗传转化及其对光合速率的调节效应 . 41(3): 507-514
ZHANG Gui-Fang, DING Zai-Song, ZHAO Ming. Transformation of Barnyardgrass ( Echinochloa crusgalli) Root Type Phosphoenolpyruvate Carboxylase Gene into Rice ( Oryza sativa) Plants and Their Effects on Photosynthesitic Gas Exchange . Acta Agronomica Sinica, 41(3): 507-514 复制到剪切板

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稗草( Echinochloa crusgalli)根型 ppc基因对水稻的遗传转化及其对光合速率的调节效应

张桂芳^1,², 丁在松¹, 赵明^1,^*

¹中国农业科学院作物科学研究所 / 农业部作物生理生态重点实验室, 北京 100081

²北京师范大学生命科学学院 / 北京师范大学学报(自然科学版)编辑部, 北京 100875

^* 通讯作者(Corresponding author): 赵明, E-mail: zhaomingcau@vip.tom.com

收稿日期:2014-12-19

基金:本研究由中国农业科学院创新工程项目资助

摘要

稗草( Echinochloa crusgalli)是稻田中的C₄光合型杂草, 为了探索稗草 ppc基因(Eppc)对水稻遗传转化的可行性及其对光合速率的调节效应, 首次将含有稗草根型磷酸烯醇式丙酮酸羧化酶(phosphoenolpyruvate carboxylase, PEPC)基因 ppccDNA的2个植物表达载体p Ubi-Eppc、p RbcS-Eppc通过农杆菌介导法对水稻进行了遗传转化。对分化植株进行的PCR、RT-PCR、克隆测序和Western杂交等结果均表明稗草 ppc基因已经整合到了水稻基因组中, 并且在转录和翻译水平都得到了表达。转基因水稻PEPC活性和气体交换参数测定结果表明T₀代多数植株的PEPC活性高于对照, 最高达到了对照的5.85倍; T₀代大多数转基因植株叶片的净光合速率( P_n)比对照提高了20.00%, 最大地提高了47.16%, 同时叶片水分利用效率(WUE)也得到增强; T₆代大部分转化植株的PEPC活性及 P_n仍保持高于对照, 本研究表明C₃根型 ppc基因过量表达也可以提高水稻的 P_n, 且证明稗草PEPC对光合作用具有较强的调节作用。

关键词: 稗草; 根型 Eppc基因; 水稻; PEPC活性; 净光合速率; 水分利用效率

Transformation of Barnyardgrass ( Echinochloa crusgalli) Root Type Phosphoenolpyruvate Carboxylase Gene into Rice ( Oryza sativa) Plants and Their Effects on Photosynthesitic Gas Exchange

ZHANG Gui-Fang^1,², DING Zai-Song¹, ZHAO Ming^1,^*

¹Institute of Crop Science, Chinese Academy of Agricultural Sciences / Key Laboratory of the Ministry of Agriculture Crop Physiology and Ecology, Beijing 100081, China

²College of Life Science / Editorial Department of Journal of Beijing Normal University (Natural Science), Beijing 100875, China

Abstract

Barnyardgrass ( Echinochloa crusgalli) is a C₄ weed commonly found in rice field. To fully utilize the photosynthestic potential of Barnyardgrass C₄gene, we transformed Barnyardgrass root Phosphoenolpyruvate Carboxylase gene into rice plant with vectors contained promoters of Ubiqitin gene and Rubisco small unit gene by Agrobactirium- mediated transformation. Both marker genes Hyg^r and ppc were detected by PCR in regenerated plants. RT-PCR and Western blot analysis confirmed that the ppc gene was incorporated into rice plant and expressed with stable transcripts and proteins. PEPC activity as measured in most of the transgenic rice plants was higher than that in control, being up to 5.85-fold of that in untransformed rice. At T₀ generation, net photosynthetic rate ( P_n) in most of transgenic rice plants was 20.00% higher than that in untransformed rice, with the highest increase of 47.16%. Water utilization efficiency (WUE) in transgenic rice was also improved. At T₆ generation, PEPC activity and P_n of transgenic lines remained higher than those of the wild type. These indicate that over-expressing C₃ Eppc gene also can improve rice photosynthesis.

Keyword: Echinochloa crusgalli; Root phosphoenolpyruvate carboxylase gene; Rice; PEPC activities; Net photosynthetic efficiency; Water use efficiency

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在C₄植物的绿色组织中, 磷酸烯醇式丙酮酸羧化酶(PEPC)是参与光合作用CO₂原初固定反应的酶。PEPC与CO₂有很强的亲和力, 其主要功能是在高温、强光下能把大气中含量很低的CO₂以四碳酸的形式固定下来, 进而运输到维管束鞘细胞叶绿体中供C₃途径利用。将C₄植物的C₄型ppc基因导入C₃作物如水稻中以提高水稻的光合能力已有多例报道^{[1, 2, 3, 4, 5]}。例如, 来源于玉米和谷子的ppc基因都被成功地导入了水稻, 在一定程度上改善了水稻的光合作用^{[1, 6]}。而稗草作为与水稻共生的主要C₄型野生植物, 比水稻具有更强的生长优势, 其ppc基因的利用还鲜有报道。在C₃植物及C₄植物的其他非光合组织中, 还存在其他形式的PEPC同工酶, 这些同工酶起着碳骨架回补作用, 参与植物的多种生理过程如气孔开放、种子发育与萌发、果实发育等^{[7, 8]}。C₃和C₄型PEPC除了具有不同的表达特征外, 其蛋白还具有不同的变构调控特征, 以适应其参与的不同生理过程。已有的研究均是将来源于C₄植物的C₄型ppc基因导入C₃植物, 而将来源于C₄植物的C₃型ppc基因导入水稻中尚未见报道。

我们前期从稗草中克隆了一个属于C₃根型的ppc基因^[9], 将该基因的cDNA分别与Ubiquitin基因启动子pUbi和1, 5二磷酸核酮糖羧化酶(Rubisco)小亚基基因启动子pRbcS融合构建了2个表达载体, 通过农杆菌介导转化中花8号水稻品种, 建立了稗草根型ppc基因(Eppc)对水稻的转化体系, 获得了一批转基因中花8号水稻植株并对其PEPC活性和气体交换特征进行了检测。

1 材料与方法

1.1 试验材料

用于遗传转化的水稻品种中花8号、菌株、质粒及载体等均由中国农业科学院作物科学研究所实验室保存; 试剂中的酶类购自TaKaRa公司; 抗生素类购自Sigma; 乙酰丁香酮购自Aldrich。

1.2 方法

1.2.1 根癌农杆菌LBA4404的转化 2个植物表达载体pUbi-Eppc和pRbcS-Eppc中, 连接稗草根型ppc基因的启动子分别为pUbi和pRbcS、终止子为nos, 采用三亲杂交法将pUbi-Eppc、pRbcS-Eppc分别转入根癌农杆菌LBA4404。

1.2.2 水稻的遗传转化水稻种子去除颖壳后, 用次氯酸钠消毒30 min; 然后接种于诱导培养基(NB培养基, 脯氨酸500 mg L^-1、谷氨酰铵500 mg L^-1、酪蛋白水解物300 mg L^-1、蔗糖30 g L^-1、2, 4-D 2 mg L^-1、植物凝胶2.6 g L^-1、pH 5.8)诱导愈伤组织生长, 每10 d继代1次。按照Toki的方法^[10]进行水稻的遗传转化。

1.2.3 GUS报告基因组织化学检测取转化处理后待分化的愈伤组织块和新分化苗(T₀代)的幼根(切段), 在新配置的GUS染色液中37℃浸泡1~2 h, 观察颜色变化情况。

1.2.4 PCR扩增和测序鉴定提取转基因T₀代植株总DNA, 对标记基因潮霉素抗性基因(Hyg^r)用引物S_hyg (5'-AAA AAG CCT GAA CTC ACC GC-3')和A_hyg (5'-ACT TCT ACA CAG CCA TCG GT-3')扩增; Eppc基因用引物P_1C(5'-ATC CGC AGA ACC CCT CCC ACT CCT CAA G-3')和P_2C(5'-GGC GTT TCT CCT CCG ACC ACT CAG CAT A-3')扩增。将扩增后的目的基因片段切胶并回收测序。

1.2.5 RT-PCR检测分别提取转基因T₀代水稻植株和非转基因中花8号(作对照)的总RNA, 进行反转录(Reverse Reaction) PCR扩增, 同样以P_1C和P_2C为扩增目的基因引物, 同时以S_actin(5'-CCC TCC CAC ATG CTA TTC T-3')和A_actin(5'-AGA GCC TCC AAT CCA GAC A-3')为引物扩增Actin基因作为内参。

1.2.6 Western杂交提取转基因T₀代水稻植株总可溶性蛋白, 通过SDS-PAGE电泳分离不同大小蛋白质, 转移至硝酸纤维素膜上依次用一抗(大豆PEPC)和二抗(碱性磷酸酶标记羊抗兔IGg)进行杂交, NBT/BCIP显色。

1.2.7 叶片PEPC活性的测定采用分光光度计法, 反应体系总体积1 mL, 组分为: 100 mmol L^-1 Tris-HCl (pH 8.0), 5 mmol L^-1 MgCl₂, 3 mmol L^-1 PEP, 0.2 mmol L^-1 NADH, 10 mmol L^-1 NaHCO₃, 10 U MDH, 以加入20 μ L酶粗提物来启动反应, 检测340 nm的吸光值下降的速率, 以单位质量可溶性蛋白来标定PEPC活性。

1.2.8 气体交换参数测定用LI-COR-6400型光合系统测定, 设定光强为1200 μ mol m^-2 s^-1, 环境温度30℃左右。分别测得净CO₂同化速率(P_n)、气孔导度(G_s)、细胞间隙CO₂浓度(C_i)、蒸腾速率(E)。叶片水分利用效率(WUE)为同步测定的P_n与E的比值。

2 结果与分析

2.1 水稻转化

接种在诱导培养基上的脱颖壳消毒后的成熟种子根伸长至3 cm左右时, 愈伤组织块在根部形成(图1-A); 转化后接种在筛选培养基上的大部分愈伤组织在短时间内发生褐化, 部分愈伤组织块约4周左右有新的黄色愈伤组织生成(图1-B); 经过抗性筛选后的愈伤组织接种至分化培养基上光照培养1周后产生绿色分化组织, 继续培养数周分化出绿色小幼苗(图1-C); 将幼苗转移到壮苗培养基上培养2周(图1-D)后成为具有根茎叶的完整水稻植株。2个载体pUbi-Eppc和pRbcS-Eppc分别获得转基因水稻植株18株和16株。

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	图1 愈伤组织诱导、转化及培养 A: 成熟胚愈伤组织诱导; B: 转化愈伤组织的筛选; C: 幼苗分化; D: 壮苗与生根。Fig. 1 Calli induction, transformation and cultivation A: calli inducement from mature embryo; B: calli transformation and selection; C: growth of regenerated plants; D: Rooting and seedling strengthening.

2.2 Gus基因表达的组织化学检测

分化前挑取愈伤组织块在GUS染色液处理后大部分愈伤组织出现了面积大小不同的蓝色晕斑; 对所获得的T₀代34株水稻苗幼根进行Gus基因组织化学检测发现: 转入2个载体pUbi-Eppc和pRbcS-Eppc的幼苗分别有16株和14株呈蓝色。表明带有目的基因的表达载体已转入水稻, 报告基因在所检测88.2%的中花8号转化苗中得到了的表达。

2.3 转基因水稻的分子鉴定

对Gus报告基因检测呈阳性的所有植株进一步的PCR鉴定发现, 30株均能扩增出约1000 bp的筛选标记基因Hyg^r的片段(图2为部分植株的PCR检测结果), 而未转基因的对照(WT: 中花8号)均无目的条带。

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图2 筛选标记基因Hyg^r的PCR检测
泳道1~7为pUbi-Eppc载体的转基因植株, 8~14为pRbcS-Eppc载体的转基因植株; M为标准DNA长度, 依次为300、500、1000、1500、2000和2500 bp, WT为未转化的品种中花8号。Fig. 2 PCR detection of Hyg resistance gene in transgenic rice
Lane 1-7: Transformants of vector pUbi-Eppc; Lane 8-14: Transformants of vector pRbcS-Eppc; M: DNA marker, being 300, 500, 1000, 1500, 2000, 2500 bp in length; WT: untransformed Zhonghua 8.

对导入的目的基因Eppc的PCR扩增发现, 30株Gus检测和Hyg^r扩增检测呈阳性的植株多数能扩增出Eppc基因0.8 kb的片段, 2个载体pUbi-Eppc和pRbcS-Eppc分别有13株和12株呈阳性(图3显示了载体pRbcS-Eppc转基因植株的PCR扩增结果), 对扩增条带回收后测序发现序列与GenBank 数据库中已登录的稗草ppc基因^[8]序列完全吻合。说明扩增出的片段的确为Eppc基因片段, 进一步证明Eppc基因已经整合到了水稻基因组内。

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	图3 转基因水稻中Eppc基因的PCR扩增检测泳道1~12为pRbcs-Eppc转基因水稻样本扩增产物, WT为对照, M为DNA标准长度, 依次为500、1000、2500、5000、7500 bp)Fig. 3 PCR detection of Eppc gene in transgenic rice Lane 1-12: Transformants of vector pRbcS-Eppc; M: DNA marker, being 500, 1000, 2500, 5000, 7500 bp in length; WT: untransformed Zhonghua 8.

在已扩增出Eppc基因的转化水稻中(pRbcS-Eppc), 随机取其中7株的RNA样本进行RT-PCR检测, 7株转基因样品Eppc基因的表达量存在差异, 且都明显比对照高, 6号样品的扩增量相对比其他样品稍低, 但高于对照; 内参基因Actin的表达量在转基因样品和对照中花8号(WT)之间基本无差别(图4)。

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	图4 转基因水稻中Eppc基因的RT-PCR检测泳道1~7为不同的转pRbcS-Eppc基因水稻植株, WT为对照中花8号。Fig. 4 RT-PCR analysis of Eppc in transgenic rice Lane 1-7: transformants of vector pRbcS-Eppc; WT: untransformed Zhonghua 8.

从转入2个表达载体pUbi-Eppc和pRbcS-Eppc的水稻植株中分别取4株进行Western杂交检测, 结果如图5。8个转化株的PEPC蛋白的表达量存在很大的差异。其中2号和6号样品比对照(WT, 中花8号)略低, 其他6个转基因样品的PEPC表达量均明显高于对照, 7号样品最高。

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	图5 转基因水稻PEPC蛋白的Western检测泳道1~4和5~8分别为pUbi-Eppc和pRbcS-Eppc载体的转基因植株, WT为对照中花8号。Fig. 5 Western analysis of PEPC in transgenic rice plants Lane 1-4: transformants of vector pUbi-Eppc; Lane 5-8: transformants of vector pRbcS-Eppc; WT: untransformed Zhonghua 8.

2.4 转基因水稻的PEPC活性

测定了25株(Eppc PCR检测为阳性)转基因水稻(T₀代)的PEPC活性(表1)。除2株(uZH8-2和uZH8-3)的PEPC活性(分别为0.0389和0.0358 μ mol mg^-1 min^-1)低于WT (0.0405 μ mol mg^-1 min^-1)外, 其余22株的PEPC活性均高于WT, 其中PEPC活性最大的1株(编号: uZH8-7)为0.2371 μ mol mg^-1 min^-1, 达到了对照的5.85倍。如图6所示的转ppc基因水稻叶片的PEPC相对活性, 2种启动子中以Ubiquitin启动子启动的ppc基因表达效率更高, 相当一部分转化株的PEPC活性比对照提高4倍以上, 而Rubisco小亚基启动子提高的活性不超过3倍。

表1 转Eppc基因水稻的PEPC活性和气体交换参数 Table 1 PEPC activity and gas exchange parameters of transgenic rice plants

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	图6 转稗草ppc基因植株的PEPC相对活性 uZH8-n: pUbi-Eppc的转化苗; rZH8-n: pRbcS-Eppc的转化苗; WT: 中花8号。Fig. 6 Relative PEPC activity improved by transgenic of Eppc in the control of different promoters uZH8-n: transformants of vector pUbi-Eppc; rZH8-n: transformants of vector pRbcS-Eppc; WT: untransformed Zhonghua 8.

2.5 转基因水稻的气体交换参数

25株T₀代转基因水稻植株的气体交换参数P_n、G_s、C_i及E都存在差别(表1); 其中4株的P_n与对照的差异小于10%, 绝大多数植株(共计15株)的P_n都比对照提高20%以上。说明PEPC活性的增加确实提高了水稻的P_n。

分析T₀代光合速率的增加幅度与PEPC活性增加程度并没有显著的相关性(图7)。由P_n/E计算得到的WUE在所有T₀代转基因植株中都得到了不同程度的增加。

表2 转Eppc基因水稻的PEPC活性及气体交换参数(T₆代) Table 2 PEPC activity and Gas exchange parameters of transgenic rice plants (T₆ generation)

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	图7 PEPC活性与净光合速率的关系缩写同图6。Fig. 7 Relationship of PEPC activities and P_n Abbreviations as the same as those given in Table 6.

经过逐代选育, 测定了10株T₆代转基因水稻的PEPC活性(表2), 发现10株的PEPC活性均高于WT, PEPC活性最大的1株达到了对照的6.676倍。相应编号T₆代花后20 d植株的气体交换参数测定结果见表2, 10株中只有1株(rZH8-12)的P_n低于对照, 其他9株的P_n均高于对照, 其中P_n最高的植株rZH8-4 (17.9 μ mol m^-2 s^-1)比对照提高了38.76%左右, 此结果证明转稗草根型ppc基因水稻在高代仍保持较高的光合速率。

3 讨论

植物的PEPC由多基因家族编码。植物的不同组织如根、茎、叶、果实、种子等器官^{[11, 12, 13]}都存在着不同类型的PEPC同工酶, 并通过组织特异性表达调节着细胞的各种生理过程。C₄光合型ppc基因主要在叶肉细胞中表达, 受光照的调控^[14], 在光合组织中主要执行原初CO₂固定的功能; 而非光合类型同工酶还具有为硝酸盐同化后的合成氨基酸提供碳骨架、维持细胞质pH和离子渗透平衡、参与种子形成与萌发、果实发育过程中物质代谢、调节气孔运动、为根瘤共生固氮过程提供碳骨架等多种功能^{[7, 15, 16, 17, 18, 19]}。

由于栽培作物玉米和谷子的C₄光合型ppc基因导入水稻后, 能使转基因水稻的光合效率不同程度地改善^{[1, 2, 3, 4, 5, 6]}, 而稗草是稻田中最常见的C₄型野生植物, 与玉米、谷子等栽培作物相比, 其更能适应水、旱2种生存环境条件, 具有更强的生长优势, 因此, 本研究选定稗草PEPC基因对水稻进行转化。根型PEPC同工酶主要在根组织表达, 根型PEPC同工酶活性的增加可提高氨的同化能力^[8]。本研究将稗草根型ppc基因分别由强组成型表达的启动子pUbi和受强光调控在叶组织特异性表达的启动子pRbcS调控下, 在转基因水稻叶组织中的表达研究发现: T₀代92%的转基因水稻P_n高于对照, 比对照提高30%以上的植株占28%; 通过逐代选育, 到T₆代检测的大部分植株P_n仍高于对照, P_n最高的植株比对照提高了38.76%左右, PEPC活性也保持高于对照。表明转稗草根型ppc基因水稻在高代大多数植株仍能保持高的光合速率, 此结果也证明C₄植物根型PEPC与叶组织表达的C₄型PEPC一样可以对C₃植物光合作用具有调节作用。国内学者凌丽俐等利用从美国华盛顿州立大学引进的转C₄光合酶基因的水稻, 经过世代繁殖、检测、选择, 得到第8代稳定的种质, 其PEPC活性经测定与第6代种质的PEPC活性相近, 他们的结果也说明PEPC基因可以稳定遗传并高表达^[20]。可见在改善C₃作物光合作用方面, 与转PPDK、NADP-ME等酶基因水稻植株的光合特性相比, PEPC起到了很关键的作用^[21], 而转PEPC+ PPDK双基因水稻植株也未表现比转PEPC基因^{[22, 23]}水稻植株有明显的光合优势^{[24, 25]}。我们转入水稻的是稗草根型PEPC基因, 我们将继续关注根型PEPC对水稻光合生理特性的改善以及在其他代谢方面的调控功能。

由气体交换参数计算得到的WUE可以反映叶片的生理抗旱能力。本研究获得的25株T₀代转基因水稻的WUE均高于对照。在玉米中导入高粱C₄型ppc基因的研究也证实WUE和抗旱能力得到了提高^[26]。我们前面的研究也已发现导入玉米和谷子C₄型ppc基因的水稻抗旱能力得到了增强^{[6, 27, 28, 29]}。本研究导入了C₄植物的根型(非光合型) ppc基因, 同样也可能提高水稻的抗旱能力, 由表1数据可知, P_n高于对照的转基因水稻中, 78.3%植株的E反而低于对照, 因而提高了WUE, 推断稗草根型PEPC同时也可能调节转基因水稻的羧化能力与气孔运动。但转基因植株的叶面积、绿叶面积的持续期及植株的干重等方面与对照没有明显差异。

转基因水稻的PEPC 活性增加的同时的确也提高了水稻的P_n, 但分析结果显示P_n增加的幅度与PEPC活性增加的程度并没有显著的相关性, 表明PEPC活性的提高被限制在一定水平内可同时提高P_n, PEPC活性的过高或过低反而对P_n的增加或降低影响不明显, 可能是与细胞质内苹果酸的代谢和运转^[30]以及苹果酸对PEPC的反馈调节作用有关^[31]。

The authors have declared that no competing interests exist.

作者已声明无竞争性利益关系。

参考文献

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1999

32.438

0.0

... 将C₄植物的C₄型ppc基因导入C₃作物如水稻中以提高水稻的光合能力已有多例报道^[1,2,3,4,5] ...

... 例如, 来源于玉米和谷子的ppc基因都被成功地导入了水稻, 在一定程度上改善了水稻的光合作用^[1,6] ...

... 由于栽培作物玉米和谷子的C₄光合型ppc基因导入水稻后, 能使转基因水稻的光合效率不同程度地改善^{[1,2,3,4,5,6]}, 而稗草是稻田中最常见的C₄型野生植物, 与玉米、谷子等栽培作物相比, 其更能适应水、旱2种生存环境条件, 具有更强的生长优势, 因此, 本研究选定稗草PEPC基因对水稻进行转化 ...

2000

4.134

0.0

... 将C₄植物的C₄型ppc基因导入C₃作物如水稻中以提高水稻的光合能力已有多例报道^[1,2,3,4,5] ...

2000

0.0

... 将C₄植物的C₄型ppc基因导入C₃作物如水稻中以提高水稻的光合能力已有多例报道^[1,2,3,4,5] ...

2002

2.922

0.0

... 将C₄植物的C₄型ppc基因导入C₃作物如水稻中以提高水稻的光合能力已有多例报道^[1,2,3,4,5] ...

2003

3.15

0.0

Fukayama

, Hatch M

, Tamai

, Tsuchida

, Sudoh

, Furbank R

, Miyao

Activity regulation and physiological impacts of maize C₄-specific phosphoenolpyruvate carboxylase overproduced in transgenic rice plants.

Photosynth Res, 2003, 77: 227-239

Phospho enol pyruvate carboxylase (PEPC) was overproduced in the leaves of rice plants by introducing the intact maize C 4 -specific PEPC gene. Maize PEPC in transgenic rice leaves underwent activity regulation through protein phosphorylation in a manner similar to endogenous rice PEPC but contrary to that occurring in maize leaves, being downregulated in the light and upregulated in the dark. Compared with untransformed rice, the level of the substrate for PEPC (phospho enol pyruvate) was slightly lower and the product (oxaloacetate) was slightly higher in transgenic rice, suggesting that maize PEPC was functioning even though it remained dephosphorylated and less active in the light. 14 CO 2 labeling experiments indicated that maize PEPC did not contribute significantly to the photosynthetic CO 2 fixation of transgenic rice plants. Rather, it slightly lowered the CO 2 assimilation rate. This effect was ascribable to the stimulation of respiration in the light, which was more marked at lower O 2 concentrations. It was concluded that overproduction of PEPC does not directly affect photosynthesis significantly but it suppresses photosynthesis indirectly by stimulating respiration in the light. We also found that while the steady-state stomatal aperture remained unaffected over a wide range of humidity, the stomatal opening under non-steady-state conditions was destabilized in transgenic rice.

1.National Institute of Agrobiological Sciences Photosynthesis Laboratory Tsukuba 305-8602 Japan 2.CSIRO Plant Industry G.P.O. Box 1600 Canberra ACT 2601 Australia

... 将C₄植物的C₄型ppc基因导入C₃作物如水稻中以提高水稻的光合能力已有多例报道^[1,2,3,4,5] ...

2013

1.051

0.0

Ding Z

, Huang S

, Zhou B

, Sun X

, Zhao

Over-expression of phosphoenolpyruvate carboxylase cDNA from C₄ millet (Seteria italica) increase rice photosynthesis and yield under upland condition but not in wetland fields.

Plant Biotechnol Rep, 2013, 7: 155-163

Phosphoenolpyruvate carboxylase (PEPC) catalyzes the initial fixation of CO2 in C-4 plants. Under the control of the rice Rubisco small subunit promoter, cDNA of a C-4 SiPPC gene cloned from Seteria italica was introduced into Japonica rice by Agrobacterium-mediated transformation. Integration of the gene was confirmed by PCR analysis. RT-PCR showed expression of the gene at the RNA level in transgenic plants, and enzyme activity measurements confirmed the increase in PEPC protein. The transformants showed improvements in both photosynthesis rate and yield only under upland field cultivation. The possible function of PEPC in rice stress tolerance is discussed.

Ding, Zai-Song 1 ;Huang, Su-Hua 1 ;Zhou, Bao-Yuan 1 ;Sun, Xue-Fang 1 ;Zhao, Ming 1 ;

... 例如, 来源于玉米和谷子的ppc基因都被成功地导入了水稻, 在一定程度上改善了水稻的光合作用^[1,6] ...

... 我们前面的研究也已发现导入玉米和谷子C₄型ppc基因的水稻抗旱能力得到了增强^[6,27,28,29] ...

2011

4.654

0.0

... 在C₃植物及C₄植物的其他非光合组织中, 还存在其他形式的PEPC同工酶, 这些同工酶起着碳骨架回补作用, 参与植物的多种生理过程如气孔开放、种子发育与萌发、果实发育等^[7,8] ...

... 而非光合类型同工酶还具有为硝酸盐同化后的合成氨基酸提供碳骨架、维持细胞质pH和离子渗透平衡、参与种子形成与萌发、果实发育过程中物质代谢、调节气孔运动、为根瘤共生固氮过程提供碳骨架等多种功能^{[7,15,16,17,18,19]} ...

2014

2.699

0.0

... 8 kb的片段, 2个载体pUbi-Eppc和pRbcS-Eppc分别有13株和12株呈阳性(图3显示了载体pRbcS-Eppc转基因植株的PCR扩增结果), 对扩增条带回收后测序发现序列与GenBank 数据库中已登录的稗草ppc基因^[8]序列完全吻合 ...

... 根型PEPC同工酶主要在根组织表达, 根型PEPC同工酶活性的增加可提高氨的同化能力^[8] ...

2005

0.0

1.667

张桂芳, 赵明, 丁在松, 张丽, 肖俊涛. 稗草磷酸烯醇式丙酮酸羧化酶(PEPCase)基因的克隆与分析. 作物学报, 2005, 31: 1365-1369

Zhang G

, Zhao

, Ding Z

, Zhang

, Xiao J

Cloning and characterization of phosphoenolpyruvate carboxylase gene from Echinochloa crusgalli.

Acta Agron Sin, 2005, 31: 1365-1369 (in Chinese with English abstract)

Phosphoenolpyruvate carboxylase (PEPCase) has a variety of functions in plants. In order to elucidate structural and functional characters of PEPCase from Echinochloa crusgalli , and search a new approach to improve crops photosynthesis, a full-length cDNA for PEPCase was isolated from E. crusgalli . Using the program of BLAST on NCBI GenBank database, the sequence presented a very high match with the genes from other plants. After alignment on ClustalW program, the identities of the cloned fragment with PEPCase genes from Setaria italica C 3 - form, Sorghum valgare C 4 - form, Zea mays C 4 - form and Oryza sativa C 3 - form were about 94.8%, 93.2%, 93.0% and 89.7%, respectively. E. crusgalli ppc ORF is 2 886 bp, encoding 961 amino acid residues. The sequence has been submitted to the GenBank database, the accession number is AY251482. Alignment and phylogenetic analysis of the amino acid sequence deduced from the fragment and the PEPCase sequences of other plants retrieved from GenBank were carried out by ClustalW program, which showed that the sequences homology of E. crusgalli with C 3 - form was higher than with C 4 - form. To identify amino acid residues and/or domains responsible for C 4 / C 3 -specific properties, we found the putative amino acid sequence contains a C 3 conserved alanine at position 771, and the sequence shared a homology of 96.5%, 96.4% with the C 3 -2- form PEPCase of Zea mays and Sorghum valgare ，83.8%, 84.3% with the C 3-1 - form PEPCase of Zea mays and Sorghum valgare ，and 82.2%,79.1%,77.1%,76.6% with the C 4 - form PEPCase of S. italica , Z. mays , S. spontaneum and Sorghum valgare . So we can conclude that the cloned sequence is a C 3 -2- form of PEPCase gene from E. crusgalli . The domain, active sites and fuction sites of Echinochloa crusgalli PEPC protein are predicted.

College of Crop Science and Bio-technology of China Agriculture University, Beijing 100094

为揭示C 4 野生植物稗草（ Echinochloa crusgalli ）PEPCase的结构和功能特点，探索改善作物高光效新途径，本研究首次克隆了稗草（ E. crusgalli ）磷酸烯醇式丙酮酸羧化酶基因( ppc )的cDNA全长，测序及同源性比较分析结果证实，稗草ppc的cDNA全长为2 886 bp,编码961个氨基酸（GenBank登录号：AY251482）；核苷酸序列与谷子( Setaria italica )C 3 型、高粱( Sorghum bicolor ) C 3-2 型、玉米( Zea mays ) C 3-2 型、水稻( Oryza sativa )C 3 型磷酸烯醇式丙酮酸羧化酶基因的同源率分别达94.8%、93.2%、93.0%和89.7%。推导的氨基酸序列中C末端第771位氨基酸是丙氨酸（A），表明是一个C 3 型基因。与其他植物几种不同形式PEPCase进行的多重序列比对与系统进化分析结果也证实，此基因的核苷酸序列及其编码的氨基酸序列与所有参与对比的C 3 型序列的同源性均远远高于与C 4 型的同源性。与玉米、高粱C 3-2 型PEPCase的同源性分别高达到96.5%、96.4%，与高粱、玉米的C 3-1 型PEPCase的同源性分别为84.3%、83.8%，而与谷子、玉米、甘蔗和高粱的C 4 型PEPCase的一致性相对较低，分别为82.2%、79.1%、77.1%、76.6%。因此进一步推论新克隆的稗草 ppc 基因属于C 3-2 型。对其编码的蛋白序列进行了结构域、活性位点和功能位点预测。

... 我们前期从稗草中克隆了一个属于C₃根型的ppc基因^[9], 将该基因的cDNA分别与Ubiquitin基因启动子pUbi和1,5二磷酸核酮糖羧化酶(Rubisco)小亚基基因启动子pRbcS融合构建了2个表达载体, 通过农杆菌介导转化中花8号水稻品种, 建立了稗草根型ppc基因(Eppc)对水稻的转化体系, 获得了一批转基因中花8号水稻植株并对其PEPC活性和气体交换特征进行了检测 ...

1997

5.319

0.0

... 按照Toki的方法^[10]进行水稻的遗传转化 ...

1990

0.0

... 植物的不同组织如根、茎、叶、果实、种子等器官^[11,12,13]都存在着不同类型的PEPC同工酶, 并通过组织特异性表达调节着细胞的各种生理过程 ...

1992

0.0

... 植物的不同组织如根、茎、叶、果实、种子等器官^[11,12,13]都存在着不同类型的PEPC同工酶, 并通过组织特异性表达调节着细胞的各种生理过程 ...

1989

3.518

0.0

... 植物的不同组织如根、茎、叶、果实、种子等器官^[11,12,13]都存在着不同类型的PEPC同工酶, 并通过组织特异性表达调节着细胞的各种生理过程 ...

1997

0.0

... C₄光合型ppc基因主要在叶肉细胞中表达, 受光照的调控^[14], 在光合组织中主要执行原初CO₂固定的功能 ...

2011

0.0

2002

3.347

0.0

Guillet

, Just

, Bénard

, Destrac-Irvine

, Baldet

, Hernould

, Causse

, Raymond

, Rothan

A fruit-specific phosphoenolpyruvate carboxylase is related to rapid growth of tomato fruit.

Planta, 2002, 214: 717-726

1.Unité de Physiologie Végétale, IBVM-INRA, BP 81, 33883 Villenave d'Ornon, France 2.Laboratoire de Biologie Cellulaire et Biotechnologie Végétales, IBVM-INRA, BP 81, 33883 Villenave d'Ornon, France 3.Unité de Recherche de Génétique et Amélioration des Fruits et Légumes (GAFL), INRA, BP 94, 84143 Montfavet, France

2004

0.0

2009

0.0

0.95

张占琴, 王金梅, 王学军, 汪凯华, 袁春新, 麻浩. 油菜籽粒发育过程中PEPCase活性与油脂、蛋白及亚基积累的特点. 中国油料作物学报, 2009, 31: 14-18

Zhang Z

, Wang J

, Wang X

, Wang K

, Yuan C

, Ma

The characteristics of PEPCase activity and accumulation of oil, protein and major protein subunits during seed development of rape (Brassica napus).

Chin J Oil Crop Sci, 2009, 31: 14-18 (in Chinese with English abstract)

以油菜品种秦优7号为材料，研究籽粒发育过程中PEPCase活性变化规律以及水份、油脂、蛋白、主要贮藏蛋白亚基等含量的积累动态，和它们之间的相关性。结果表明，种子发育期间，水份含量从81.61%下降到23.22%，油脂含量从0.26%上升到31.38%，蛋白含量从1.1%上升到21.36%，可溶性蛋白含量从0.63%上升到12.21%。12S球蛋白先积累，2S清蛋白后积累；PEPCase活性与蛋白含量、油脂含量分别呈阶段性相关，与油脂含量/蛋白含量比值呈极显著负相关；与12S组分含量呈显著负相关，与α 2-3，α 4，β2，β3，β4亚基含量呈显著负相关，与其它亚基含量的相关性不显著；与2S组分及其亚基含量相关性不显著。

2013

0.0

Pan L

, Yang Q

, Chi X

, Chen M

, Yang

, Chen

, Wang

, He Y

, Yu S

Functional analysis of the phosphoenolpyruvate carboxylase on the lipid accumulation of peanut (Arachis hypogaea L. ) seeds.

J Integr Agric, 2013, 12: 36-44

Phosphoenolpyruvate carboxylase (PEPC; EC 4.1.1.31) catalyses phosphoenolpyruvate (PEP) to yield oxaloacetate, which is involved in protein biosynthesis. Pyruvate kinase (PK; EC 2.7.1.40) catalyzes PEP to yield pyruvate, which is involved in fatty acid synthesis. In this study, five PEPC genes (AhPEPC1, AhPEPC2, AhPEPC3, AhPEPC4, and AhPEPC5) from peanut have been cloned. Using a quantitative real-time RT-PCR approach, the expression pattern of each gene was monitored during the seed development of four peanut varieties (E11, Hebeigaoyou, Naihan 1, and Huayu 26). It was found that these five genes shared similar expression behaviors over the developmental stages of E11 with high expression levels at 30 and 40 d after pegging (DAP); whereas these five genes showed irregular expression patterns during the seed development of Hebeigaoyou. In Naihan 1 and Huayu 26, the expression levels of the five genes remained relatively high in the first stage. The PEPC activity was monitored during the seed development of four peanut varieties and seed oil content was also characterized during whole period of seed development. The PEPC activity followed the oil accumulation pattern during the early stages of development but they showed a significantly negative correlation thereafter. These results suggested that PEPC may play an important role in lipid accumulation during the seed development of four peanut varieties tested.

Shandong Peanut Research Institute, Qingdao 266100, P.R.China

2006

0.0

1.667

凌丽俐, 林宏辉, 焦德茂. 转PEPC基因水稻种质的稳定光合生理特性. 作物学报, 2006, 32: 527-531

Ling L

, Lin H

, Jiao D

The stable photosynthetic characteristics of a PEPC transgenic rice germplasm.

Acta Agron Sin, 2006, 32: 527-531 (in Chinese with English abstract)

The eighth generation of PEPC transgenic rice and an untransformed rice cultivar ( Oryza sativa L. subsp. japonica cv. Kitaake) were need to study photosynthetic rate and chlorophyll fluorescence parameters by applying the stable and radioactive carbon isotope. The results showed that the germplasm of PEPC transgenic rice was a C 3 plant with certain C 4 photosynthetic characteristics.The light saturation photosynthetic rate of transgenic rice was higher than that of untransformed rice. In the treatment with high light intensity (3 h) and methyl viologen (MV), a photooxidative reagent, the PSⅡ photochemical efficiency (F v /F m ) and photochemical quenching (q P ) of transgenic PEPC rice were decreased less, while the non-photochemical quenching (qN) were increased more as comparison with those of untransformed rice. The higher activities of antioxidative enzymes superoxide dismutase (SOD) and peroxidase (POD) in transgenic rice plants led to a lesser accumulation of endogenous active oxygen (O 2 ．- , H 2 O 2 ). As a result, the malondialdehyde (MDA, a product of membrane-lipid oxidation) content was reduced. Compared with the previous generations, the germplasm of PEPC transgenic rice has been steady in genetic, which may become a new germplasm resource in the breeding for high photosynthetic efficiency.

Key Laboratory of Bio-resources and Eco-environment, Ministry of Education, College of Life Science, Sichuan University, Chengdu 610064, Sichuan

本文运用稳定和放射性碳同位素研究了第8代转PEPC基因水稻，并测定其光合速率和荧光特性，结果表明该种质为C 3 植物，但C 4 原初光合产物增多，表现出C 4 光合特性有所改善；转PEPC基因水稻种质已稳定，为我国的高光效育种提供了一个新的种质资源。

... 国内学者凌丽俐等利用从美国华盛顿州立大学引进的转C₄光合酶基因的水稻, 经过世代繁殖、检测、选择, 得到第8代稳定的种质, 其PEPC活性经测定与第6代种质的PEPC活性相近, 他们的结果也说明PEPC基因可以稳定遗传并高表达^[20] ...

0.0

... 可见在改善C₃作物光合作用方面, 与转PPDK、NADP-ME等酶基因水稻植株的光合特性相比, PEPC起到了很关键的作用^[21], 而转PEPC+ PPDK双基因水稻植株也未表现比转PEPC基因^[22,23]水稻植株有明显的光合优势^[24,25] ...

2001

0.0

0.95

焦德茂, 李霞, 黄雪清, 迟伟, 匡廷云, 古森本. 转PEPC基因水稻的光合CO₂同化和叶绿素荧光特性. 科学通报, 2001, 46: 414-418

Jiao D

, Li

, Huang X

, Chi

, Kuang T

, Gu S

The characteristics of CO₂ assimilation of photosynthesis and chlorophyll fluorescence in transgenic PEPC rice.

Chin Sci Bull, 2001, 46: 414-418

以转 PEPC,PPDK,NADP-ME,PEPC+PPDK 基因水稻及原种为材料,比较C 4 途径有关光合酶活性、叶绿素荧光参数、CO 2 交换等生理指标,重点研究了转 PEPC 基因水稻的生理特性, 结果如下:(1)转 PEPC 基因水稻 PEPC 活性比原种高20倍,光饱和光合速率比原种高55%,羧化效率提高50%,CO 2 补偿点降低27%;(2)在高光强(3 h)或光氧化剂甲基紫精(MV)处理后, 与原种相比,转 PEPC 基因水稻的PSⅡ光化学效率( F v / F m )、光化学猝灭( q P )下降较少,证明其耐光抑制和耐光氧化能力增强;(3)在高光强条件下,转 PEPC 基因水稻中RuBPCase活性变化不明显,但碳酸酐酶(CA)诱导活性增加1.8倍.这些结果为揭示高光合效率机理和高光合效率育种提供了有益的启示.

2003

0.0

2002

0.599

0.0

2008

0.0

1.889

张边江, 华春, 周峰, 周泉澄, 陈全战, 王荣富, 焦德茂. 转PEPC+PPDK双基因水稻的光合特性. 中国农业科学, 2008, 41: 3008-3014 (in Chinese with English abstract)

Zhang B

, Hua

, Zhou

, Zhou Q

, Chen Q

, Wang R

, Jiao D

Photosynthetic characteristics of transgenic rice with PEPC+PPDK gene.

Sci Agric Sin, 2008, 41: 3008-3014 (in Chinese with English abstract)

安徽农业大学生命科学学院

【目的】系统研究ATP处理后转PEPC+PPDK双基因水稻的光合特性,证明ATP是增强转C4基因水稻光合能力的关键因子。【方法】以原种和转PEPC+PPDK双基因水稻为材料，进行了PCR检测，C4光合酶活性的测定。通过ATP处理后，分析了光、温—光合曲线和活性氧代谢有关指标，统计分析了相关的产量构成因素。【结果】原种中虽有全套的C4光合酶，但活性很低。PCR检测出玉米的PEPC和PPDK基因转入普通水稻后，转PEPC+PPDK双基因水稻高表达了C4光合酶活性。在高光和高温条件下，同未施ATP的相比, ATP处理后转PEPC+PPDK双基因水稻光合速率分别提高17%和12%。在光氧化条件下，耐光氧化能力进一步增强，产量提高15%。【结论】ATP处理后,转PEPC+PPDK双基因水稻增强了光合生产力,表明ATP是设计类似C4水稻的关键因子。

2002

3.142

0.0

Jeanneau

, Gerentes

, Foueillassar

, Zivy

, Vidal

, Toppan

, Perez

Improvement of drought tolerance in maize: towards the functional validation of the Zm-Asr1 gene and increase of water use efficiency by over-expressing C₄-PEPC.

Biochimie, 2002, 84: 1127-1135

Abstract Water availability is one of the major limiting factors for plant growth. Maize is particularly sensitive to water stress at reproductive stages with a strong impairment of photosynthesis and grain filling. Here, we describe the use of genetic transformation first to assess the role of a candidate gene Asr1—a putative transcription factor—as an explanation for genetically linked drought tolerance Quantitative Trait Loci (QTLs), and second to modify CO 2 fixation rates in leaves through changes of C 4 phosphoenolpyruvate carboxylase (C 4 -PEPC) activity. Transgenic Asr1 over-expressing lines show an increase in foliar senescence under drought conditions. The highest C 4 -PEPC overexpressing line exhibited an increase (+30%) in intrinsic water use efficiency (WUE) accompanied by a dry weight increase (+20%) under moderate drought conditions. Opposite effects were observed for transgenic plants under-expressing the corresponding proteins. The data presented here indicate the feasibility to increase the level of endogenous biochemical activities related to water economy and/or drought tolerance, and opens a way to develop maize varieties more tolerant to dry growing conditions.

... 在玉米中导入高粱C₄型ppc基因的研究也证实WUE和抗旱能力得到了提高^[26] ...

2007

0.0

1.667

丁在松, 赵明, 荆玉祥, 李良璧, 匡廷云. 玉米ppc基因过表达对转基因水稻光合速率的影响. 作物学报, 2007, 33: 717-722

Ding Z

, Zhao

, Jing Y

, Li L

, Kuang T

Effect of overexpression of maize ppc gene on photosynthesis in transgenic rice plants.

Acta Agron Sin, 2007, 33: 717-722 (in Chinese with English abstract)

Overexpression of ppc gene in transgenic rice to improve photosynthesis and yield have brought extensive interest in the world. Many attempts have been made to transfer maize ppc gene to rice plants using genetic engineering techniques. But there was still a controversy on whether overexpression of PEPC could increase photosynthetic rates (P n ) in transgenic rice plants. We thought the different results were caused by the difference on materials, ppc gene inserting site and the measuremental circumstance. In order to validate our ideas, we repeated to introduce maize ppc gene into rice plants by Agrobacterium -mediated transformation and a lot of transgenic rice plants were obtained. PCR, Southern, Western analyse and PEPC activity detection indicated that the maize ppc gene was integrated into rice genome and expressed correctly and efficiently. The Pn was 27.3% higher in one transgenic line with the highest PEPC activity than in untransformed plants. The increase is at least in part due to the enhanced stomatal conductivity and higher internal CO 2 concentration. Nearly all the ppc transgenic rice lines showed higher Pn (up to 68.8%) than untransformed plants grown in dry field and full sunlight. Compared with that in appropriated conditions, the 6 transgenic lines showed higher Pn in stress condition. These results indicated that photosynthesis can be improved by introduction of PEPC into rice, especially in stress conditions.

1 Key Laboratory of Photosynthesis and Environmental Molecular Physiology, Institute of Botany, Chinese Academy of Sciences, Beijing 100093; 2 Graduate School of Chinese Academy of Sciences, Beijing 100049; 3 Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing 100081, China

通过转基因技术将外源ppc基因导入水稻以期增加产量是国内外的重要研究领域。然而，关于ppc基因过表达能否提高转基因水稻的光合速率至今尚无定论。本研究将玉米ppc基因导入水稻中花8号，获得大量转基因水稻植株。经PCR筛选、PEPC活性测定、Southern和Western杂交分析，表明玉米ppc基因已经整合到受体水稻基因组中，并得到了正确和高效的表达。测定了温室内种植的T 1 代6个PEPC活性不同的转基因水稻植株的光合速率，只有活性最高的株系的光合速率显著增加，增加的幅度为27.3%。在旱作栽培条件下测定了T 3 代33个转基因株系的光合速率，结果表明在高温、高光强下，绝大多数转基因水稻的光合速率增加，最大提高了68.8%。比较6个转基因株系的T 1 和T 3 代看到，逆境胁迫条件下转基因水稻光合速率明显提高。以上结果表明水稻中导入ppc基因可以提高其光合速率，特别是逆境条件下的光合速率。

... 我们前面的研究也已发现导入玉米和谷子C₄型ppc基因的水稻抗旱能力得到了增强^[6,27,28,29] ...

2008

0.0

1.667

方立锋, 丁在松, 赵明. 转ppc基因水稻苗期抗旱特性研究. 作物学报, 2008, 34: 1220-1226

Fang L

, Ding Z

, Zhao

Characteristics of drought tolerance in ppc overexpressed rice seedlings.

Acta Agron Sin, 2008, 34: 1220-1226 (in Chinese with English abstract)

Introducing enzymes involved in C 4 photosynthesis into C 3 crops is supposed to enhance the photosynthesis and crop productivity. Several attempts have been made to transfer phosphoenolpyruvate carboxylase (PEPC) gene from maize into rice, however, the relation of photosynthetic capacity, productivity and stress tolerance in transgenic rice was not analyzed. In the present research, the productivity and photosynthesis in T 4 offspring of ppc gene overexpressed rice were investigated under drought stress. The results indicated that the yield increased by 28% and 42%, and the number of tillers increased by 27% and 40%, respectively, in two transgenic lines T1 and T2 compared with those of the wild type under upland cultivation. Under drought stress, the transgenic rice seedlings maintained higher photosynthesis (A), stomatal conductance ( G s ) and water use efficiency (WUE), as well as higher superoxide dismutase (SOD) activity and lower malondialdehyde (MDA) content than the wild type. It is suggested that the overexpression of ppc gene in rice makes higher antioxidation ability under drought stress. In addition, the transgenic lines T1 and T2 exhibited higher proline content and lower increase of osmotic potential under drought stress, suggesting that the ppc gene transgenic rice has the higher osmoregulation ability than the wild type. The similar results were also found in the ex-periments treated by PEG-6000. Therefore, the present results suggested that, the ppc gene overexpressed rice confers higher photosynthesis and drought tolerance under drought stress, which is helpful to increase the number of tillers, then contribute to enhance the rice yield.

Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing 100081, China

通过C 4 转基因技术改善C 3 作物光合作用, 以期提高作物产量是国内外研究的热点之一。然而, 目前关于转磷酸烯醇式丙酮酸羧化酶(PEPC)基因对水稻的光合作用、产量和抗旱性的影响及其调节机理仍不很清楚。本研究以T 4 代转 ppc 基因水稻为材料, 进行产量和苗期抗旱性研究。结果表明, 在旱作栽培条件下, 两个转ppc基因株系(T1, T2)单株产量分别比未转基因的对照(WT)增产28%和42%, 分蘖增加27%和40%; T1和T2可维持较高的光合速率、气孔导度和水分利用效率; 干旱胁迫使超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量增加, T1和T2 SOD活性增幅(+25%)都显著高于WT(+9%), 而MDA含量增幅显著低于WT, 表明转ppc基因水稻具有较强的抗氧化能力; T1和T2比WT具有较高的脯氨酸含量和渗透势下降幅度, 说明转PEPC基因水稻的渗透调节能力高于对照。PEG-6000处理下, 得到的结果相似。

... 我们前面的研究也已发现导入玉米和谷子C₄型ppc基因的水稻抗旱能力得到了增强^[6,27,28,29] ...

2011

0.0

1.667

周宝元, 丁在松, 赵明. PEPC过表达可以减轻干旱胁迫对水稻光合的抑制作用. 作物学报, 2011, 37: 112-118

Zhou B

, Ding Z

, Zhao

Alleviation of drought stress inhibition on photosynthesis by overexpression of PEPC gene in rice.

Acta Agron Sin, 2011, 37: 112-118 (in Chinese with English abstract)

Introducingenzymes involved in photosynthesis of C 4 plants into rice is supposed to enhance the photosynthesis and crop productivity. However, only a few researches showed that the photosynthesis and crop productivity has been improved by introducing phosphoenolpyruvate carboxylase (PEPC) gene into rice. In the present research, the photosynthesis in 42 lines of PEPC gene overexpressed rice was investigated. The average photosynthetic rate ( P n ) of transgenic rice lines was almost the same as that of the wild type (control) in paddy field, while much higher in upland field. Only a few transgenic lines showed higher P n in paddy field and most of them showed higher P n in upland field. Similar results were found in the water controlled experiment. Two transgenic rice lines with different relative activities of PEPC (10 and 25 fold) were selected to study their photosynthesis under different water potentials (0, -20, and -40 kPa). In both lines, P n was similar with that in the wild type under normal condition (0 kPa) and much higher under drought conditions (-20 and -40 kPa). In both experiments, the transgenic lines had higher P n under drought conditions, with a much slower decreasing rate than the wild type. Therefore, the present results suggested that the overexpressed PEPC could not improve the photosynthetic rate of transgenic rice plants. But the photosynthetic rate of transgenic rices declined slowly under drought condition. So it is supposed that PEPC might be involved in drought resistance to decrease the inhibition of drought stress on photosynthesis in rice.

Institute of Crop Sciences, Chinese Academy of Agricultural Sciences / Key Laboratory of Crop Ecophysiology and Cultivition, Ministry of Agriculture, Beijing 100081, China

为了明确磷酸烯醇式丙酮酸羧化酶(PEPC)过量表达能否提高水稻的光合速率，测定了42个表达不同PEPC水平的转玉米PEPC基因水稻株系及对照(受体亲本中花8号)开花期和灌浆期的光合速率。结果表明，在水田条件下，转基因株系光合速率与未转基因对照相比没有明显差异；而在旱地条件下，转基因水稻的光合速率显著高于对照(27%和24%)。随机选取2个PEPC相对活性分别为10倍和25倍的转基因株系进行网室精确控水盆栽实验得到相似的结果。说明单纯导入PEPC并不能提高水稻的光合速率，而干旱胁迫下转基因水稻的光合优势可能是由于PEPC参与水稻的抗旱反应而减轻了干旱胁迫对光合作用的抑制作用。

... 我们前面的研究也已发现导入玉米和谷子C₄型ppc基因的水稻抗旱能力得到了增强^[6,27,28,29] ...

2004

6.555

0.0

... 转基因水稻的PEPC 活性增加的同时的确也提高了水稻的P_n, 但分析结果显示P_n增加的幅度与PEPC活性增加的程度并没有显著的相关性, 表明PEPC活性的提高被限制在一定水平内可同时提高P_n, PEPC活性的过高或过低反而对P_n的增加或降低影响不明显, 可能是与细胞质内苹果酸的代谢和运转^[30]以及苹果酸对PEPC的反馈调节作用有关^[31] ...

1987

3.582

0.0