Plastid transformation in Arabidopsis thaliana

拟南芥质体转化

• 批准号: 1716102
• 负责人: Pal Maliga
• 金额: $ 62万
• 依托单位: Rutgers University New Brunswick
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1716102&HistoricalAwards=false
• 关键词: Plastid; transformation; Arabidopsis; thaliana

Engineering the chloroplast genome is key to improving the efficiency of photosynthesis, the process that converts sunlight energy into biomass in crops. In most plants, chloroplast engineering is very inefficient. However, a recent breakthrough has revealed the genetic bottleneck for this process, and this project will leverage the new information to develop methods for efficient engineering of chloroplasts in the model plant, Arabidopsis thaliana. The experimental design will be directly applicable to crops in the evolutionary related Brassicaceae family, broccoli, an important vegetable, and oilseed rape, an important source of edible oil. The ability to engineer chloroplasts in these crops could lead to significant production improvements. The project will also have educational impact by providing research training for undergraduates, including women and underrepresented minorities. Undergraduates will be recruited from Rutgers University through The Douglass Project for Rutgers Women in Math, Science and Engineering, and The Rutgers Aresty Research Center for Undergraduates. Under-represented minorities, low-income, and first-generation college students will also be recruited from the Collegiate Science and Technology Entry Program or the First in the World Grant programs at Farmingdale State College, Farmingdale, NY. A summer research experience in chloroplast biotechnology will be offered to an undergraduate student from the University of Sao Paulo (USP), Brazil as part of a plan to recruit students for the joint Rutgers/USP PhD program. Transformation of the plastid genome is routine in tobacco, but 100-fold less frequent in the model plant, Arabidopsis thaliana, preventing its widespread use for studying questions in plastid biology. Recent data have revealed that plastid transformation efficiency is 100-fold enhanced in Arabidopsis plants with a defective ACC2 gene. This mutant line is hypersensitive to spectinomycin, the selective agent used for plastid transformation. Proof of principle for increased plastid transformation frequency was obtained in the Columbia background, because ACC2 T-DNA knockout lines are available in this accession. However, the Columbia accession is recalcitrant to plant regeneration from cultured cells where plastid transformation is carried out. The goal of this research is to provide Arabidopsis plastid transformation research tools for the plant community. This will be achieved through (1) obtaining ACC2 knockout lines in the regenerable RLD and Ws accessions; (2) developing plant regeneration protocols from transplastomic cells to obtain fertile transplastomic plants; (3) obtaining regenerable forms of the Columbia ecotype; and (4) constructing plastid transformation vectors that are suitable to achieve these goals. The results will accelerate studies of plastid function, thereby allowing better understanding of the 'rules' that govern interactions between this important organelle and the nucleus and contribute to essential plastid functions.This award is co-funded by the Genetic Mechanisms Program in the Division of Molecular and Cellular Biosciences and by the Plant Genome Research Program in the Division of Integrative Organismal Systems; both programs are in the Directorate for Biological Sciences.

改造叶绿体基因组是提高光合作用效率的关键，光合作用是将光能转化为作物生物量的过程。在大多数植物中，叶绿体工程是非常低效的。 然而，最近的一项突破揭示了这一过程的遗传瓶颈，该项目将利用新的信息开发在模式植物拟南芥中有效改造叶绿体的方法。 该实验设计将直接适用于进化相关的油菜科作物，花椰菜，一种重要的蔬菜，和油菜，一种重要的食用油来源。 在这些作物中设计叶绿体的能力可能会导致显着的产量提高。该项目还将通过为大学生，包括妇女和代表性不足的少数民族提供研究培训，产生教育影响。罗格斯大学的本科生将通过罗格斯大学数学、科学和工程专业的道格拉斯项目以及罗格斯大学的阿雷斯蒂研究中心招募。代表性不足的少数族裔、低收入和第一代大学生也将从纽约州法明代尔州立学院的大学科学技术入学计划或世界第一助学金计划中招募。叶绿体生物技术的夏季研究经验将提供给巴西圣保罗大学（USP）的一名本科生，作为罗格斯大学/USP博士联合项目招生计划的一部分。质体基因组的转化在烟草中是常规的，但在模式植物拟南芥中的频率要低100倍，这阻止了其广泛用于研究质体生物学问题。最近的数据显示，在具有缺陷ACC 2基因的拟南芥植物中，质体转化效率提高了100倍。该突变株系对用于质体转化的选择剂大观霉素高度敏感。在哥伦比亚背景中获得了质体转化频率增加的原理证明，因为ACC 2 T-DNA敲除系在该登录中可用。然而，哥伦比亚的加入不利于从进行质体转化的培养细胞再生植物。本研究的目的是为植物界提供拟南芥质体转化的研究工具。这将通过（1）在可再生的RLD和Ws种质中获得ACC 2敲除系;（2）从转质体细胞开发植物再生方案以获得可育的转质体植物;（3）获得哥伦比亚生态型的可再生形式;和（4）构建适于实现这些目标的质体转化载体来实现。这一成果将加速对质体功能的研究，从而更好地理解这一重要细胞器与细胞核之间相互作用的“规则”，并有助于质体的基本功能。该奖项由分子和细胞生物科学部遗传机制计划和综合有机体系统部植物基因组研究计划共同资助;这两个项目都在生物科学局。

项目成果

Prospects for Reengineering Agrobacterium tumefaciens for T-DNA Delivery to Chloroplasts

重组根癌农杆菌将 T-DNA 递送至叶绿体的前景

• DOI: 10.1093/plphys/kiab081
• 发表时间: 2021
• 期刊: Plant Physiology
• 影响因子: 7.4
• 作者: Matsuoka, Aki;Maliga, Pal
• 通讯作者: Maliga, Pal

New Tools for Engineering the Arabidopsis Plastid Genome

工程拟南芥质体基因组的新工具

• DOI: 10.1104/pp.19.00761
• 发表时间: 2019
• 期刊: Plant Physiology
• 影响因子: 7.4
• 作者: Yu, Qiguo;LaManna, Lisa;Kelly, Megan E.;Lutz, Kerry Ann;Maliga, Pal
• 通讯作者: Maliga, Pal