PlantTransform: TRTech-PGR: Genotype-independent Regeneration for Recalcitrant Species Through Induced Totipotent Plant Cells

生物技术在作物性状改良方面的应用为实施更可持续和更具弹性的农业实践带来了巨大的希望。例如，经过改良的作物需要更少的水和肥料补充，或者更耐旱和更耐植物病害的作物的产生，可以帮助提高作物产量，以应对气候变化和人口规模的增加。尽管我们在单个细胞或组织中修改植物基因组(编码作物的可遗传性状)的能力已经取得了长足的进步，但将这些细胞或组织重新转变为完整的植物仍然是生物技术应用于作物性状改良的一个重大瓶颈。该项目的目的是寻找有效的方法对细胞进行重新编程，以促进它们转化和再生为完整的植物。为此，我们将在模式植物系统thale cress中进行研究，以更好地了解再生的细胞和分子调控，并确定可以促进这一过程的因素。作为原则的证明，我们随后将在一种已知难以转化和再生的重要作物品种--小麦--中使用这些因素。除了计划中的研究，我们还将通过为弗吉尼亚州夏季住宅州长农业学校的高中生开发一系列讲座来促进公众的理解和青年的参与，题为《作物性状改良的生物工程的过去、现在和未来》。再生是植物转化的瓶颈，我们对控制再生的分子调控网络知之甚少。该项目汇集了一个多学科的研究团队，以开发能够推进植物转化领域的技术，并进一步了解细胞全能的基本方面。重点将是通过异源表达形态转录因子来提高再生效率，重点是从原生质体培养中再生。通过原生质体瞬时转化的基因组编辑技术的应用，为快速产生无转基因编辑植株，特别是在高度杂合、异交或营养繁殖的作物物种中提供了巨大的希望。该项目利用拟南芥作为模型来研究原生质体培养的再生。我们将使用自动数字图像分析和发育中的微愈伤组织的单细胞转录图谱作为一个高通量、信息丰富的平台。通过常规的体细胞组织培养和原生质体培养，将对20个形态发生转录因子在拟南芥中对再生的影响进行比较筛选。我们将研究由促进原生质体培养再生的转录因子控制的基因调控网络，以从机制上深入了解它们的功能。有效的转录因子将在冬小麦中进行测试，冬小麦是一种抵抗转化和组织培养的物种。这个项目将促进我们对全能性和细胞命运决定的基础性理解。这个奖项反映了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。

The application of biotechnology for crop trait improvement holds great promise for the implementation of more sustainable and resilient agricultural practices. For instance, the generation of crops that have been modified to require less water and fertilizer supplementation or that are more tolerant of drought conditions and resistant to plant diseases can help improve crop yields in the face of a changing climate and increasing population size. Although we have made great strides in our ability to modify the plant genome (that encodes the heritable traits of our crops) in individual cells or tissues, turning those cells or tissues back into whole plants presents a significant bottleneck in the application of biotechnology for crop trait improvement. The aim of this project is to find effective ways to reprogram cells in order to facilitate their transformation and regeneration into whole plants. To that end, we will perform studies in a model plant system, thale cress, to better understand the cellular and molecular regulation of regeneration and identify factors that can enhance the process. As proof of principle, we will subsequently implement the use of such factors in an important crop species known to be difficult to transform and regenerate, namely wheat. In addition to the planned research, we will also advance public understanding and youth involvement through the development of a lecture series for high school students in the Virginia Summer Residential Governor's School for Agriculture titled The Past, Present, and Future of Bioengineering for Crop Trait Improvement. Regeneration is a bottleneck for plant transformation, and we know little about the molecular regulatory networks that govern regeneration. This project brings together a multi-disciplinary research team to develop technology that can advance the field of plant transformation and further understanding the fundamental aspects of cell totipotence. The focus will be to enhance regeneration efficiency through ectopic expression of morphogenic transcription factors, with an emphasis on regeneration from protoplast culture. The application of genome editing technology through transient transformation of protoplasts holds great promise for the rapid generation of transgene-free edited plants, especially in highly heterozygous, outcrossing, or vegetatively propagated crop species. The project utilizes Arabidopsis as a model to investigate regeneration from protoplast culture. We will use automated digital image analysis and single-cell transcript profiling of developing microcalli as a high-throughput, information-rich platform. A collection of 20 morphogenic transcription factors will be comparatively screened in Arabidopsis for their effects on regeneration through conventional somatic tissue culture and protoplast culture. The gene regulatory networks governed by transcription factors that promote regeneration from protoplast culture will be studied to give mechanistic insight into their function. Effective transcription factors will be tested in winter wheat, a species recalcitrant to transformation and tissue culture. This project will advance our foundational understanding of totipotency and cell fate determination.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.