2种耐干藓类再生体系优化及遗传转化体系构建

Desiccation tolerance (DT) means the organs could endure almost totally water loss, but could recover within seconds encounter water. It's one kind of extremely water deficit tolerance in the nature, and the plant which possess DT traits is also called "the resurrection plants". Vegetative desiccation tolerance only existed in the alga, lichen, bryophytes, and very small part of angiosperms. Because of the complicated structural protection and cellular repairment during the course of desiccation tolerance, the vegetative desiccation tolerance mechanism, taken bryophytes as target, had gained more and more attention. Based on the past research, we had targeted 2 moss survived in desert, Syntrichia caninervis and Bryum Argenteum, which show excellent desiccation tolerance trait in the profile of ecology and physiology. In the research, we committed to build up the genetic transformation system of the two mosses. Firstly, gametophyte regeneration system of S. caninervis should be optimized, by testing the culture component, culture condition, and matching the hormone ratio; secondly, genetic transformation method should be studied, by using GFP reporter genes, and groping toward the suitable detection method for transgenetic shoots; Thirdly, genetic transformation system should be validated by instill verified functional genes, DREB and ALDH21 genes. Upon above research, the practicability and manipuility of two system should be evaluated, based on the index of protonema differentiation rate, homologous recombination rate, genetic transformation rate and degree of tissue cultural difficulty. Finally, the potential bryophytes desiccation tolerance model should be put forward. The research will build fundamental basis for further research on the new gene function involved in the DT, and will help to shed light on the molecular mechanism of desiccation tolerance.

齿肋赤藓和银叶真藓在极端干旱时组织完全脱水,但"干而不死"，并遇水快速复苏，是自然界难得的研究耐干胁迫的好材料。为了深入研究其耐干机制,迫切需要建立配子体再生体系及遗传转化体系，以实现大量单克隆材料的获取及原位分子遗传学操作。本项目拟优化2种藓类配子体再生体系并构建遗传转化体系。通过齿肋赤藓再生体系优化（外植体筛选、培养条件、激素配比筛选）、2种藓类遗传体系建立（载体构建、遗传转化方法、原生质体获得与再生培养）、遗传转化体系验证（过表达与基因敲除）等三个方面研究，建立稳定、高效的再生及遗传转化体系，并基于生活周期、培养难易、分子操作的角度筛选，确立最佳的耐干藓类模式种。项目的顺利实施，可为大量耐干相关的（新）基因功能原位验证（基因打靶技术）提供平台，有助于发现功能强大新基因；此外，2种藓类是沙漠生物结皮的优势种，研究可为沙漠"生物地毯"工程的实施提供技术支撑以及源源不断的生物材料。

齿肋赤藓和银叶真藓在极端干旱时组织完全脱水,但”干而不死”，并遇水快速复苏，是自然界难得的研究耐干胁迫的好材料。为了深入研究其耐干机制,迫切需要建立配子体再生体系及遗传转化体系，以实现大量单克隆材料的获取及原位分子遗传学操作。本项目优化了2种藓类配子体再生体系并构建了其瞬时遗传转化技术体系。通过齿肋赤藓再生体系优化（外植体筛选、培养条件、激素配比筛选）、2种藓类瞬时遗传转化体系建立（载体构建、遗传转化方法与再生培养）、遗传转化体系验证（GUS基因及DREB抗性基因过表达）等三个方面研究，建立稳定、高效的再生及瞬时遗传转化体系。此外，研究了2种耐干藓类银叶真藓和齿肋赤藓对非生物胁迫（干、盐、高温）的抗性生理，探究了不同耐干（快速干燥、缓慢干燥、大气干燥）过程相对含水量的变化，并深入研究了2种材料不同高温梯度胁迫下（80℃,100℃,110℃,120℃）形态、超微结构、生理生化及抗性基因的表达变化，并首次发现植物界耐高温的世界纪录，即银叶真藓可耐受120℃20分钟。此外，对采自新疆境内17个居群300余个个体的银叶真藓表型可塑性及遗传多样性进行了研究，并探讨了其形态/遗传变异与环境可塑性的关系。项目的顺利实施，一方面为大量耐干相关的（新）基因功能原位验证（基因打靶技术）提供平台，有助于发现功能强大新基因；另一方面，2种藓类是沙漠生物结皮的优势种，研究可为沙漠“生物地毯”工程的实施提供技术支撑以及源源不断的生物材料，具有较为重要的开发应用潜力。