Development of Agrobacterium-mediated transient expression system in Caragana intermedia and characterization of CiDREB1C in stress response
Development of Agrobacterium-mediated transient expression system in Caragana intermedia and characterization of CiDREB1C in stress response
复制标题
农杆菌介导的中间柠条瞬时表达系统的开发以及 CiDREB1C 在胁迫反应中的表征
DOI：
10.1186/s12870-019-1800-4
复制
发表时间：
2019-06-06
影响因子：
5.3
通讯作者：
Li, Guojing
中科院分区：
文献类型：
--
作者：
Liu, Kun;Yang, Qi;Li, Guojing
背景：农杆菌介导的瞬时转化是快速分析植物基因功能的一种通用且不可缺少的方法。尽管这种瞬时表达系统已经成功地应用于许多植物物种，但它在锦鸡儿中发育不全。结果：本研究建立了农杆菌介导的中间叶瞬时表达体系，并优化了不同农杆菌菌株、几种表面活性剂和Silwet L-77浓度对中间叶瞬时表达效率的影响。在5株农杆菌中，GV3101的GUS表达量最高。此外，Silwet L-77的瞬时表达量高于Triton X-100和Tween-20。0.001%浓度的Silwet L-77显著提高了GUS瞬时表达水平。Real-time PCR结果显示，瞬时表达植株中CiDREB1C的表达量大幅上调，并在浸润后第2天达到最高水平。基于此优化的瞬态转化方法，研究了CiDREB1C对干旱、盐和ABA处理的响应。结果表明，在干旱、盐和ABA处理下，与对照苗相比，瞬态表达CiDREB1C能提高中叶参成活率和叶绿素含量，降低倒伏率。此外，ABA处理下CiDREB1C瞬时表达苗的叶片脱落率低于对照。结论：以菌株GV3101和浓度为0.001%的Silwet L-77混合农杆菌侵染中叶，获得了最佳的瞬时表达条件。瞬时表达的CiDREB1C增强了对干旱、盐和ABA胁迫的耐受性，这表明它是一种合适而有效的工具，可以用来确定与中间体非生物胁迫响应相关的基因功能。
Background:The Agrobacterium-mediated transient transformation is a versatile and indispensable way of rapid analyzing gene function in plants. Despite this transient expression system has been successfully applied in a number of plant species, it is poorly developed in Caragana intermedia.Results:In this study, we established an Agrobacterium-mediated transient expression system in C. intermedia leaves and optimized the effect of different Agrobacterial strains, several surfactants and the concentration of Silwet L-77, which would affect transient expression efficiency. Among the 5 Agrobacterial strains examined, GV3101 produced the highest GUS expression level. Besides, higher level of transient expression was observed in plants infiltrated with Silwet L-77 than with Triton X-100 or Tween-20. Silwet L-77 at a concentration of 0.001% greatly improved the level of GUS transient expression. Real-time PCR showed that expression of CiDREB1C was highly up-regulated in transiently expressed plants and reached the highest level at the 2nd day after infiltration. Based on this optimized transient transformation method, we characterized CiDREB1C function in response to drought, salt and ABA treatment. The results showed that transiently expressed CiDREB1C in C. intermedia leaves could enhance the survival rate and chlorophyll content, and reduce the lodging rate compared with the control seedlings under drought, salt and ABA treatments. Furthermore, the rate of leaf shedding of CiDREB1C transient expression seedlings was lower than that of the control under ABA treatment.Conclusions:The optimized transient expression condition in C. intermedia leaves were infiltrated with Agrobacterial strains GV3101 plus Silwet L-77 at a concentration of 0.001% added into the infiltration medium. Transiently expressed CiDREB1C enhanced drought, salt and ABA stress tolerance, indicated that it was a suitable and effective tool to determine gene function involved in abiotic stress response in C. intermedia.