Monte-Bello Carolina C, Araujo Elias F, Martins Marina C M, Mafra Valeria, da Silva Viviane C H, Celente Viviane, Caldana Camila
Brazilian Bioethanol Science and Technology Laboratory (CTBE), Brazilian Center for Research in energy and materials (CNPEM); University of Campinas (UNICAMP).
Brazilian Bioethanol Science and Technology Laboratory (CTBE), Brazilian Center for Research in energy and materials (CNPEM); University of Viçosa (UFV).
J Vis Exp. 2018 Aug 25(138):57800. doi: 10.3791/57800.
A wide range of studies in plant biology are performed using hydroponic cultures. In this work, an in vitro hydroponic growth system designed for assessing plant responses to chemicals and other substances of interest is presented. This system is highly efficient in obtaining homogeneous and healthy seedlings of the C3 and C4 model species Arabidopsis thaliana and Setaria viridis, respectively. The sterile cultivation avoids algae and microorganism contamination, which are known limiting factors for plant normal growth and development in hydroponics. In addition, this system is scalable, enabling the harvest of plant material on a large scale with minor mechanical damage, as well as the harvest of individual parts of a plant if desired. A detailed protocol demonstrating that this system has an easy and low-cost assembly, as it uses pipette racks as the main platform for growing plants, is provided. The feasibility of this system was validated using Arabidopsis seedlings to assess the effect of the drug AZD-8055, a chemical inhibitor of the target of rapamycin (TOR) kinase. TOR inhibition was efficiently detected as early as 30 min after an AZD-8055 treatment in roots and shoots. Furthermore, AZD-8055-treated plants displayed the expected starch-excess phenotype. We proposed this hydroponic system as an ideal method for plant researchers aiming to monitor the action of plant inducers or inhibitors, as well as to assess metabolic fluxes using isotope-labeling compounds which, in general, requires the use of expensive reagents.
植物生物学领域的众多研究都采用水培法进行。在本研究中,我们展示了一种体外水培生长系统,该系统旨在评估植物对化学物质及其他感兴趣物质的反应。该系统能高效地分别培育出C3和C4模式植物拟南芥和绿色狗尾草的均匀且健康的幼苗。无菌培养避免了藻类和微生物污染,而这是水培中已知的限制植物正常生长发育的因素。此外,该系统具有可扩展性,能够大规模收获植物材料且机械损伤较小,如有需要还能收获植物的各个部分。本文提供了详细的实验方案,证明该系统组装简便且成本低廉,因为它使用移液器架作为植物生长的主要平台。我们使用拟南芥幼苗评估雷帕霉素靶蛋白(TOR)激酶的化学抑制剂AZD - 8055的效果,验证了该系统的可行性。在AZD - 8055处理后30分钟,就能在根和芽中有效检测到TOR抑制作用。此外,经AZD - 8055处理的植物表现出预期的淀粉过量表型。我们认为这种水培系统是植物研究人员的理想方法,可用于监测植物诱导剂或抑制剂的作用,以及使用通常需要昂贵试剂的同位素标记化合物来评估代谢通量。