Zhang Lijie, Fu Jingqi, Dong Tianyi, Zhang Mengmeng, Wu Jingwen, Liu Chunping
Key Laboratory of Forest Tree Genetics, Breeding and Cultivation of Liaoning Province, Shenyang Agricultural University, Shenyang, China.
Key Laboratory of Silviculture of Liaoning Province, Shenyang Agricultural University, Shenyang, China.
Front Plant Sci. 2023 Oct 12;14:1243030. doi: 10.3389/fpls.2023.1243030. eCollection 2023.
(Manchurian walnut) is a precious timber and woody grain and oil species in Northeast China. The heterodichogamous characteristic phenomenon resulted in the non-synchronous flowering and development of male and female flowers, which limited the mating and the yield and quality of fruits. is a core gene in the flowering regulatory networks, which has been cloned in , and the function has also been verified preliminarily. In this study, the promoter sequence with different lengths of 5'-deletion (pLFY1-pLFY6) were cloned and conducted bioinformatics analysis, the promoter activities were analyzed by detecting their driving activity to GUS gene in the tobacco plants that transformed with different promoter sequence stably or transiently. After that, the interaction between JmSOC1 and gene promoter was also analyzed via yeast single-hybrid. The results showed that the promoter sequence contains core cis-acting elements essential for eukaryotic promoters, hormone response elements, defense- and stress-responsive elements, flowering-related elements, etc. Transgenic tobacco plants with were obtained by infection using the pCAMBIA1301 expression vector, and the GUS gene driven by the promoter was detected to express in the leaf, stem, flower, and root of the transformed tobacco plant, which indicated that the obtained promoter had driving activity. GUS histochemical staining and enzyme activity detection showed that promoter fragments with different lengths had promoter activity and could respond to the induction of long photoperiod, low temperature, salicylic acid (SA), IAA, GA3, and methyl jasmonate (MeJA). The core regulatory region of gene promoter in was between -657 bp and -1,904 bp. Point-to-point validation of yeast single-hybrid confirmed the interaction between JmSOC1 and gene promoter, which indicated that gene is the downstream target of JmSOC1. These results reveal relevant factors affecting gene expression and clarify the molecular mechanism of gene regulation in the flower developmental partially, which will provide a theoretical basis for regulating the flowering time by regulating gene expression in .
（胡桃楸）是中国东北地区珍贵的用材及木本粮油树种。其雌雄异熟特性导致雌雄花开花及发育不同步，限制了授粉及果实产量和品质。LFY是开花调控网络中的核心基因，已在[具体物种]中克隆，功能也已初步验证。本研究克隆了不同长度5'-缺失的LFY启动子序列（pLFY1-pLFY6）并进行生物信息学分析，通过检测其对稳定或瞬时转化不同启动子序列的烟草植株中GUS基因的驱动活性来分析启动子活性。之后，还通过酵母单杂交分析了JmSOC1与LFY基因启动子之间的相互作用。结果表明，该启动子序列包含真核生物启动子必需的核心顺式作用元件、激素响应元件、防御和胁迫响应元件、开花相关元件等。利用pCAMBIA1301表达载体通过农杆菌介导法获得了转LFY基因的烟草植株，检测到LFY启动子驱动的GUS基因在转化烟草植株的叶、茎、花和根中表达，表明获得的LFY启动子具有驱动活性。GUS组织化学染色和酶活性检测表明，不同长度的启动子片段均具有启动子活性，且能响应长日照、低温、水杨酸（SA）、吲哚乙酸（IAA）、赤霉素（GA3）和茉莉酸甲酯（MeJA）的诱导。胡桃楸LFY基因启动子的核心调控区域在-657 bp至-1904 bp之间。酵母单杂交点对点验证证实了JmSOC1与LFY基因启动子之间的相互作用，表明LFY基因是JmSOC1的下游靶标。这些结果揭示了影响LFY基因表达的相关因素，部分阐明了胡桃楸花发育过程中LFY基因调控的分子机制，将为通过调控胡桃楸LFY基因表达来调控开花时间提供理论依据。
