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中的应用为实施更具可持续性和抗灾能力的农业实践带来了巨大的希望。例如，在气候变化和人口规模增加的情况下，培育需要更少的水和肥料补充或更能忍受干旱条件和抵抗植物疾病的作物有助于提高作物产量。尽管我们在修改单个细胞或组织中的植物基因组（对作物的可遗传性状进行编码）的能力方面取得了很大的进步，但在将这些细胞或组织转化为整个植物的过程中，生物技术在作物性状改善方面的应用遇到了一个重大瓶颈。该项目的目的是找到有效的方法来重编程细胞，以促进其转化和再生成整个植物。为此，我们将在一个模式植物系统中进行研究，芥蓝，以更好地了解再生的细胞和分子调节，并确定可以增强这一过程的因素。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，即小麦中实施这些因子的使用。除了计划中的研究之外，我们还将通过在弗吉尼亚州夏季住宿州长农业学校为高中生开发一个名为“作物性状改良生物工程的过去、现在和未来”的系列讲座，促进公众的理解和青少年的参与。再生是植物转化的瓶颈，而我们对调控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，以开发技术，可以推进植物转化领域和进一步了解细胞全能的基本方面。重点将是通过形态发生转录因子的异位表达来提高再生效率，重点是原生质体培养再生。原生质体瞬时转化基因组编辑技术的应用为快速生成无转基因编辑植物提供了巨大的希望，特别是在高杂合、异交或无性繁殖的作物物种中。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单细胞转录分析来开发微愈伤组织，作为一个高通量，信息丰富的平台。通过常规体细胞组织培养和原生质体培养，比较筛选20个形态发生转录因子对拟南芥再生的影响。由转录因子控制的促进原生质体再生的基因调控网络将被研究，以提供对其功能的机制见解。有效的转录因子将在冬小麦中进行试验，这是一种难以转化和组织培养的物种。这个项目将推进我们对全能性和细胞命运决定的基本理解。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。

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.