Plant Transformation Facility, Donald Danforth Plant Science Center, Saint Louis, Missouri.
Curr Protoc. 2021 May;1(5):e127. doi: 10.1002/cpz1.127.
Setaria viridis is an emerging model system for the genetic and molecular characterization of cereals and bioenergy crops. Here, we describe a detailed procedure for genetic transformation of the S. viridis accession line ME034V-1. This method utilizes callus generated from mature seeds for infection with Agrobacterium tumefaciens strain AGL1 to regenerate hygromycin-resistant stable transgenic plants. It takes approximately 7 weeks to generate callus from mature seeds, 11-17 weeks from infection to the regeneration of transgenic lines, and an additional 3-4 weeks for plant growth in the greenhouse for seed collection. The protocol as presented consistently results in transformation frequency of approximately 25% for the generation of transgenic plants, with fewer escapes and higher survivability in soil for optimal seed collection. © 2021 Donald Danforth Plant Science Center. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Generation of S. viridis (Accession ME034V-1) callus from mature seeds Basic Protocol 2: Agrobacterium-mediated transformation of callus to generate transgenic plants Basic Protocol 3: Plantlet transplantation in soil, plant growth in greenhouse, and seed collection Support Protocol: Preparation of Agrobacterium culture for infection.
狗尾草是一种新兴的模式系统，可用于研究谷物和生物能源作物的遗传和分子特性。本文描述了狗尾草 ME034V-1 品系的遗传转化的详细程序。该方法利用成熟种子产生的愈伤组织，用根癌农杆菌 AGL1 菌株感染，以再生潮霉素抗性稳定的转基因植株。从成熟种子中产生愈伤组织大约需要 7 周，从感染到转基因系再生需要 11-17 周，在温室中生长 3-4 周以收集种子。按照本方案进行操作，可稳定获得约 25%的转基因植株转化频率，用于优化种子收集的土壤中逃脱的植株较少，存活率更高。© 2021 唐纳德丹佛斯植物科学中心。当前协议由 Wiley Periodicals LLC 出版。基本方案 1：从成熟种子中生成狗尾草（ME034V-1 品系）愈伤组织基本方案 2：用根癌农杆菌介导的愈伤组织转化生成转基因植株基本方案 3：在土壤中移植植物苗、在温室中生长植物并收集种子支持方案：用于感染的根癌农杆菌培养物的制备。
