CAREER: Elucidating trans-kingdom horizontal gene transfer mechanisms to improve plant genetic engineering

植物是气候危机许多潜在解决方案的核心。然而，开发具有新的有用功能的植物可能需要数年时间。植物转化和负面舆论被广泛认为是培育植物新品种的主要瓶颈。该团队将推行一项综合研究和教育计划，以促进植物基因工程的创新。这项研究将产生与农杆菌和植物之间跨王国DNA转移有关的基础知识，这些知识将被用于开发植物转化工具，以提高在植物中实现的遗传操作的速度和复杂性。同时，PI将为注册参加其国家教育公平实验室课程的学生开发一个暑期计划：BIOE80“生物工程导论”。PI已经与国家教育公平实验室合作-这是一个非营利性组织，为低收入(标题1)高中生提供机会，通过向大学教授学习大学课程来获得免费大学学分。拟议的夏季BIOE80课程将使学生有机会获得在以讲座为基础的课程中学到的基因工程技术的实践经验。它将在学生对生物工程概念的理论理解和在STEM领域进行实习和高等教育所需的研究经验之间架起亟需的桥梁。最终，这门课程和拟议的研究应该会加强美国植物工程的论述，研究应该有助于更充分地实现植物生物技术对可持续未来的潜力。跨王国将DNA从农杆菌转移到植物是一项惊人的生物学壮举，也是强大的植物生物技术工具的基础。尽管是将新的DNA导入植物最常用的工具，但在我们对农杆菌介导的DNA转移的理解中仍然存在一些重要的差距-包括转移的DNA(T-DNA)整合到植物基因组中的动力学和机制。这一知识差距阻碍了可用于可靠地控制转基因在植物基因组中插入位置的植物基因组工程工具的产生，并限制了在植物中实现遗传扰动的速度和复杂性。这项研究将研究在两种最常见的植物转化过程：花浸渍和愈伤组织体外接合后，T-DNA在植物细胞中传递和T-DNA命运(整合或降解)的时间动态。该团队将使用PI开发的一种新的合成遗传系统，该系统可以记录T-DNA的传递，以前所未有的精度剖析这些复杂的生物过程。然后，新获得的信息将被用于创建分子工具，用于在没有T-DNA整合的情况下进行植物基因组编辑。这项工作将通过产生开发日益复杂的植物基因组工程工具所需的基础知识，并为精确设计植物以提高气候适应性的长期目标奠定基础，从而为生物经济做出贡献。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Plants lie at the heart of many potential solutions to the climate crisis. Yet, it can take years to develop plants with new, useful features. Plant transformation and negative public opinion are widely recognized as the main bottlenecks to engineering new plant varieties. This team will pursue an integrated research and education plan to spur innovation in plant genetic engineering. The research will generate foundational knowledge related trans-kingdom DNA transfer between Agrobacterium and plants, which will be used to develop plant transformation tools that improve the speed and complexity of genetic manipulation achievable in plants. In parallel, the PI will develop a summer program for students enrolled in their National Education Equity Lab course: BIOE80 “Introduction to Bioengineering.” The PI already works with the National Education Equity lab – a non-profit that provides low-income (Title 1) high school students with an opportunity to earn free college credits by taking college courses from college professors. The proposed summer addition to BIOE80 will give students an opportunity to gain hands-on experience with the genetic engineering techniques learned in their lecture-based course. It will serve as much needed bridge between the students’ theoretical understanding of bioengineering concepts and the research experience needed to pursue internships and higher education in STEM field. Ultimately, the course and proposed research should strengthen U.S. plant engineering discourse and research should help more fully realize the potential of plant biotechnology for a sustainable future.Trans-kingdom transfer of DNA from Agrobacterium to plants is a stunning biological feat and the basis of powerful plant biotechnology tools. Despite being the most commonly used tool for introducing new DNA to plants, several important gaps in our understanding of Agrobacterium-mediated DNA transfer remain – including the dynamics and mechanisms of transferred DNA (T-DNA) integration into the plant genome. This knowledge gap has prevented the generation of plant genome engineering tools that can be used to reliably control the insertion location of transgenes in plants’ genomes and limits the speed and complexity of genetic perturbation achievable in plants. The proposed research will investigate the temporal dynamics of T-DNA delivery and T-DNA fate (integration or degradation) in plant cells after the two most common plant transformation procedures: floral dip and in vitro conjugation to callus. The team will use a new synthetic genetic system, developed by the PI, that can record T-DNA delivery to dissect these complex biological processes with unprecedented precision. Then, newly acquired information will be used to create molecular tools for plant genome editing without T-DNA integration. This work will contribute to the bioeconomy by generating the foundational knowledge needed to develop increasingly sophisticated plant genome engineering tools and setting the stage for the long-term goal of precisely engineering plants in order to improve climate resilience.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.