过表达胡杨PeCPK7提高拟南芥耐盐性

Overexpression of Populus euphratica PeCPK7 enhancing salt tolerance in Arabidopsis thaliana

【目的】研究胡杨PeCPK7在植物耐盐过程中的作用,旨在进一步揭示植物耐盐的生理与分子调控机制。【方法】根据NCBI基因组数据库中胡杨PeCPK7 CDS序列,克隆PeCPK7基因,利用DNAMAN进行氨基酸序列比对,并用Mega 7软件进行进化树构建。以过表达PeCPK7拟南芥(PeCPK7-OE1、PeCPK7-OE2和PeCPK7-OE3)、野生型(WT)和转空载体对照(VC)拟南芥株系为试验材料,对各基因型拟南芥进行不同盐浓度处理,从生理生化和分子生物学水平研究胡杨PeCPK7在盐胁迫中的响应机制。【结果】(1)PeCPK7蛋白与其他物种中的CPK7有高度相似性,并与毛果杨PtrCPK7家族亲缘关系最近。(2)盐处理胡杨幼苗,PeCPK7的相对表达量在盐胁迫6 h后达到最大值,至48 h时恢复至最初状态。(3)PeCPK7定位于细胞质中。(4)在盐处理后,过表达PeCPK7拟南芥株系的生存率和根长均显著高于野生型(WT)和转空载体(VC)拟南芥;并且根细胞膜受损伤程度显著低于WT和VC。(5)在根中积累的Na^(+)和H_(2)O_(2)显著低于WT和VC,Ca^(2+)显著高于WT和VC;并且Na^(+)外排和K^(+)内流显著高于WT和VC。(6)盐胁迫下,过表达株系的超氧化物歧化酶、过氧化物酶和过氧化氢酶活性升高幅度显著高于WT和VC。(7)过表达株系的叶绿素含量、PSⅡ最大光量子效率、实际光合量子产量、相对电子传递速率和光合速率高于WT和VC,而胞间CO_(2)浓度低于WT和VC。【结论】过表达胡杨PeCPK7能够提高拟南芥的耐盐能力,本研究将为利用基因工程提高植物的耐盐性提供理论依据。

[Objective]This study aimed to reveal the physiological and molecular regulation mechanisms of PeCPK7 in plant salt stress tolerance.[Method]According to the CDS of PeCPK7 in NCBI genome database,the PeCPK7 was cloned,DNAMAN was used for amino acid sequence alignment,and Mega 7 software was used for evolutionary tree construction.The mechanism of PeCPK7 in salt stress was studied from the level of physiology,biochemistry and molecular biology by Arabidopsis thaliana overexpressed lines(PeCPK7-OE1,PeCPK7-OE2 and PeCPK7-OE3),the wild-type(WT)and vector control(VC)as experimental materials.[Result](1)PeCPK7 was highly similar to CPK7 in other species,and was closely related to PtrCPK7 family of Populus trichocarpa.(2)In salt-treated Populus euphratica seedlings,the relative expression of PeCPK7 reached the maximum after 6 h of salt stress,and recovered to the initial state at 48 h.(3)PeCPK7 was localized in the cytoplasm.(4)After salt treatment,the survival rate and root length of overexpression lines were significantly higher than those of WT and VC.(5)Na^(+)and H_(2)O_(2) accumulated in the root were significantly lower than those of WT and VC,and Ca^(2+)was significantly higher than WT and VC.The Na^(+)efflux and K^(+)influx were significantly higher than WT and VC.(6)Under salt stress,the activity of antioxidant enzymes(SOD,POD,CAT)of overexpression lines were significantly higher than WT and VC.(7)The decrease of chlorophyll content,PSII.maximum optical quantum efficiency,actual photosynthetic quantum yield,relative electron transfer rate and the photosynthetic rates of overexpression lines were higher than those of WT and VC,while the intercellular CO_(2) concentration was lower than WT and VC.[Conclusion]Overexpression of PeCPK7 gene can improve salt stress tolerance of Arabidopsis thaliana,which will provide a theoretical basis for improving plant salt tolerance through genetic engineering.