油茶节律基因LHY的克隆及其在外源激素处理下对光合作用的影响  

Cloning of circadian clock gene LHY and its effect on photosynthesis under exogenous hormone treatment in Camellia oleifera

作　　者：何之龙 张震[1] 许彦明[1] 王瑞[1] 王湘南[1] 陈永忠[1] HE Zhilong;ZHANG Zhen;XU Yanming;WANG Rui;WANG Xiangnan;CHEN Yongzhong(Hunan Academy of Forestry,Changsha 410004,Hunan,China)

机构地区：[1]湖南省林业科学院,湖南长沙410004

出　　处：《经济林研究》2021年第4期1-9,共9页Non-wood Forest Research

基　　金：湖南省重点研发计划项目“油茶‘双高’、宜机新种质创制与繁育技术研究”(2020NK2050);湖南省自然科学基金项目“节律基因表达调控油茶光合作用的机制研究”(2019JJ50303)。

摘　　要：【目的】为提高油茶光合效率的高光效定向育种研究提供新思路。【方法】以油茶主推良种‘湘林210’为试验材料,基于油茶叶片转录组测序数据设计特异性引物,利用RT-PCR技术克隆油茶LHY基因的全长序列,并对克隆的目的片段序列及其编码的氨基酸序列开展生物信息学分析。在人工气候箱中模拟中日照光周期,对试验材料喷施100μg/L的ABA和100 mg/L的GA溶液,研究不同激素处理对油茶苗木光合作用指标和节律基因表达水平的影响。【结果】克隆获得了油茶节律基因LHY目的片段共2565 bp,预测开放阅读框为2301 bp,编码766个氨基酸,其编码蛋白的相对分子质量为83.88 kD。通过对油茶LHY基因编码氨基酸序列的分析,预测油茶LHY基因编码蛋白为亲水性的不稳定酸性蛋白质,其二级结构以无规则卷曲结构为主。油茶LHY基因编码蛋白的氨基酸序列经同源比对和系统发育树分析,结果表明其与茶树LHY基因编码的氨基酸序列同源性最高,相似性为91.06%。GA和ABA信号均对油茶幼苗的光合作用生理指标和节律基因表达有显著调控作用。100 mg/L的GA处理在中日照光周期中能显著提高净光合速率、气孔导度和蒸腾速率,并在一定程度上促进LHY基因表达;100μg/L的ABA处理在中日照光周期中能显著抑制净光合速率和蒸腾速率,并在一定程度上抑制LHY基因表达。【结论】成功克隆了油茶节律基因LHY全场编码区序列,并且发现油茶幼苗光合作用指标对不同植物激素的响应与LHY基因表达调控趋势一致。【Objective】Providing the high photosynthetic efficiency directional breeding studies with new thoughts in Camellia oleifera.【Method】The main recommended variety of C.oleifera,‘Xianglin 210’,was used as experimental material.Specific primers were designed based on transcriptome sequencing data of C.oleifera leaves,the full-length sequence of LHY gene was cloned by RT-PCR,and bioinformatics analysis was carried out on the obtained target fragment and its encoding amino acid sequence.In order to study the effects of different plant hormone treatments on photosynthesis index and circadian clock gene expression level of C.oleifera seedlings,100 μg/L ABA and 100 mg/L GA solution were sprayed on the experimental materials in an artificial climate chamber to simulate the mid-day photoperiod.【Result】The target fragment of LHY gene in C.oleifera consisted 2565 bp,its predicted open reading frame consisted 2301 bp,encoding 766 amino acids,of which relative molecular weight was 83.88 kd.By analyzing the amino acid sequence encoded by LHY gene in C.oleifera,it was predicted that LHY protein in C.oleifera was a hydrophilic and unstable acidic protein,and its secondary structure was mainly random coil structure.The amino acid sequence encoded by LHY gene in C.oleifera was analyzed by homology alignment and phylogenetic tree.The results showed that the amino acid sequence encoded by LHY gene in C.oleifera had the highest homology with the amino acid sequence encoded by LHY gene in C.sinensis,and the similarity was 91.06%.The results showed that GA and ABA could significantly regulate the photosynthetic parameters and circadian gene expression in C.oleifera seedlings.100 mg/L GA significantly increased net photosynthetic rate,stomatal conductance and transpiration rate,as well as LHY gene expression to a certain extent;contrarily,100μg/L ABA treatment significantly decreased net photosynthetic rate and transpiration rate,meanwhile inhibited LHY gene expression to a certain extent.【Conclusion】The coding seque

关 键 词：油茶 节律基因 植物激素 基因表达 

分 类 号：S601[农业科学—园艺学]