摘要
【目的】对巨桉Eucalyptus grandis中特有的低温响应基因EgrCIN1 (Eucgr.B02882)序列及其表达特征进行分析,并探索其在植物低温响应中发挥的功能,以丰富桉树抗寒基因资源。【方法】利用生物信息学手段分析EgrCIN1基因、蛋白序列的特征和启动子上的顺式作用元件信息;采用实时荧光定量PCR (RT-qPCR)技术分析EgrCIN1在巨桉不同组织及4℃不同时间、干旱、 300 mmol·L^(-1)1-氯化钠(NaCl)、 100μmol·L^(-1)脱落酸(ABA)和100μmol·L^(-1)茉莉酸甲酯(MeJA)处理下植株叶片中的表达特征;通过EgrCIN1::GFP载体瞬时转化烟草Nicotiana tabacum进行亚细胞定位;并构建CaMV35S启动子驱动的过表达载体异源转化拟南芥Arabidopsis thaliana,获得3个转基因纯合株系后,对其进行-6℃低温、0.5μmol·L^(-1)ABA等逆境处理,分析EgrCIN1在低温胁迫响应中发挥的功能。【结果】EgrCIN1是巨桉中特有的基因,不含内含子,没有跨膜结构,启动子含有多个与逆境响应相关的顺式作用元件。该基因在茎和叶片中都有表达,而在根中不表达;在叶片中其表达受低温处理强烈诱导;同时,干旱、高盐和ABA等非生物逆境因子也能诱导叶片中EgrCIN1的表达,其编码蛋白被定位在叶绿体中。拟南芥中EgrCIN1过表达转基因株系对低温耐受性提高,对外源ABA敏感程度增强。【结论】EgrCIN1是在叶绿体表达,并可能通过ABA依赖途径参与植物低温胁迫响应,提高植物对低温的抗性。图8表2参27。
[Objective] The objective is to analyze the sequence and expression characteristics of EgrCIN1(Eucgr.B02882), a unique low temperature response gene in Eucalyptus grandis, and to explore its function in plant low temperature response, so as to enrich gene resources to cold resistance. [Method] Bioinformatics methods were used to analyze the characteristics of EgrCIN1 gene, protein sequence and cis-acting elements on the promoter. The expression pattern of EgrCIN1 in different tissues, at 4 ℃ in different time, under drought condition, under treatment of 300 mmol·L^(-1)NaCl, 100 μmol·L^(-1)ABA and 100 μmol·L^(-1)MeJA were analyzed by RT-qPCR method. Subcellular localization of EgrCIN1 was completed by transient transformation of Nicotiana tabacum with EgrCIN1::GFP vector. Besides, an overexpression vector driven by CaMV35S promoter was constructed and transformed into Arabidopsis thaliana. Three transgenic lines were obtained and treated with low temperature(-6 ℃) and ABA(0.5 μmol·L^(-1)) to analyze the function of EgrCIN1 in response to low temperature stress. [Result] EgrCIN1 was a unique gene induced by low temperature in E. grandis. It did not contain introns and had no transmembrane structure. The promoter contained multiple cis acting elements related to stress response. The gene was mainly expressed in leaves and stems, but not in roots.Moreover, its expression in leaves was strongly induced by low temperature treatment. Meanwhile, abiotic stress factors such as drought, high salt and ABA could also induce EgrCIN1 expression in leaves. The encoded protein of EgrCIN1 was localized in chloroplasts. In Arabidopsis, EgrCIN1 overexpression transgenic lines were more tolerant to low temperature and more sensitive to exogenous ABA. [Conclusion] EgrCIN1 is expressed in chloroplasts and may participate in plant cold stress response possibly through ABA-dependent pathway to improve plant cold resistance. [Ch, 8 fig. 2 tab. 27 ref.]
作者
李芳燕
夏晓雪
吴梦洁
洪家都
程龙军
LI Fangyan;XIA Xiaoxue;WU Mengjie;HONG Jiadu;CHENG Longjun(College of Forestry and Biotechnology,Zhejiang A&F University,Hangzhou 311300,Zhejiang,China)
出处
《浙江农林大学学报》
CAS
CSCD
北大核心
2022年第6期1194-1202,共9页
Journal of Zhejiang A&F University
基金
浙江省科技厅重大科技专项(2021C02070-6)。
