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

植物是许多气候危机潜在解决方案的核心。然而，培育出具有新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植物新品种工程的主要瓶颈。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。该研究将产生有关农杆菌和植物之间跨界DNA转移的基础知识，这些知识将用于开发植物转化工具，以提高植物基因操作的速度和复杂性。与此同时，PI将为参加国家教育公平实验室课程的学生开发一个暑期项目：BIOE80“生物工程导论”。PI已经与国家教育公平实验室（National Education Equity lab）合作。国家教育公平实验室是一家非营利机构，为低收入（Title 1）高中学生提供通过学习大学教授的大学课程来获得免费大学学分的机会。拟议的夏季增加到BIOE80将给学生一个机会，以获得实践经验与遗传工程技术在他们的讲座为基础的课程学习。它将成为学生对生物工程概念的理论理解与追求STEM领域实习和高等教育所需的研究经验之间急需的桥梁。最终，该课程和拟议的研究应该加强美国植物工程的论述和研究应该帮助更充分地实现植物生物技术的潜力，以实现可持续的未来。从农杆菌到植物的DNA跨界转移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。尽管农杆菌是向植物引入新DNA的最常用工具，但我们对农杆菌介导的DNA转移的理解仍然存在一些重要的空白，包括转移DNA （T-DNA）整合到植物基因组的动力学和机制。这种知识差距阻碍了植物基因组工程工具的产生，这些工具可用于可靠地控制转基因在植物基因组中的插入位置，并限制了植物中可实现的遗传扰动的速度和复杂性。该研究将研究在两种最常见的植物转化过程：花浸渍和离体接合到愈伤组织后，T-DNA传递和T-DNA命运（整合或降解）在植物细胞中的时间动态。该团队将使用由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，以前所未有的精度解剖这些复杂的生物过程。然后，新获得的信息将用于创建不需要T-DNA整合的植物基因组编辑分子工具。这项工作将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，并为精确改造植物以提高气候适应能力的长期目标奠定基础，从而为生物经济做出贡献。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值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.