耐旱大豆品种中GmAUX1基因对干旱响应的分子调控机制及激素信号途径研究

Searching for the key genes for drought tolerance from drought resistant cultivars is an effective way to improve soybean drought tolerance and cultivate new varieties of drought tolerance. Previous work showed that under drought, GmAUX1 gene was specifically expressed in drought-tolerant soybean variety, and it may be a downstream gene of MYB transcription factor. However, whether the GmAUX1 gene can improve drought tolerance of soybean and the molecular mechanism of drought tolerance are not clear. In the study, the GmAUX1 gene expression changes under auxin or ABA treatment, subcellular localization, the analysis of promoter differences between the drought-tolerant and drought-sensitive varieties, whether the GmAUX1 gene is directly regulated by MYB transcription factor, the selection of interaction proteins of GmAUX1, the influence of the target gene expression to soybean in response to drought, the observation of root morphology and structure of transgenic soybean lines and the detection of the downstream genes expression of GmAUX1 gene, are reveal the molecular mechanism of soybean GmAUX1 gene involved in drought response, which can provide theoretical and technical support for the genetic improvement of soybean drought tolerance, and has important significance for improving soybean yield and quality.

寻找耐旱品种中干旱响应的关键基因是提高大豆耐旱能力，培育耐旱新品种的有效途径。申请者研究发现，干旱条件下，GmAUX1基因在耐旱大豆品种中特异表达，并且在MYB基因下游表达。但该基因在大豆中的耐旱性及耐旱分子机制尚不明确。本项目以GmAUX1基因作为研究对象，研究生长素及脱落酸处理对该基因表达的影响及GmAUX1基因的亚细胞定位；分析耐旱与不耐旱品种间目标基因启动子差异并寻找耐旱品种中目标基因响应干旱的核心启动子区域与元件；通过酵母单（双）杂交系统验证目标基因是否受MYB直接调控，并筛选目标基因的互作蛋白；通过超表达及RNA沉默GmAUX1基因，研究目标基因的表达对大豆耐旱性的影响；观测转基因大豆的根部形态与结构；分析目标基因在生长素通路中下游基因的表达。研究结果将揭示大豆GmAUX1基因参与干旱响应的分子调控机理，为大豆耐旱性遗传改良提供理论与技术支撑，对改良大豆产量与品质具有重要意义。

寻找耐旱品种中干旱响应的关键基因是提高大豆耐旱能力，培育耐旱新品种的有效途径。申请者在项目实施期间，研究发现GmLAXs-I (GmAUX1) 基因主要定位于细胞膜，GmLAXs-I表达受脱落酸和生长素响应，并且在PEG、干旱胁迫处理后的表达量下调，推测GmLAXs-I是干旱胁迫的负调控因子。GmLAX4在耐旱品种与干旱敏感品种中的启动子序列存在SNPs。通过构建Cas9基因编辑载体，转化大豆，获得了lax1/3、lax4、lax9/15突变体，并初步验证了lax4突变体比野生型的耐旱性增强。通过免疫共沉淀技术及高通量测序分析（chip-seq），表明GmLAX3可能是MYB转录因子（GmMYB14）调控的靶基因。研究结果初步明确了部分GmLAXs基因耐旱的生物学功能及上游调控作用因子，其他GmLAXs基因的耐旱性继续验证。