宁夏枸杞LbALDH3F1基因克隆及耐盐性分析  

Cloning of LbALDH3F1 Gene from Lycium barbarum and Analysis of Its Salt Tolerance

作　　者：马小霞 胡进红 梁旺利 麻玉荣 孙雯 王玲霞[1,2] 秦垦 梁文裕[1,2] MA Xiaoxia;HU Jinhong;LIANG Wangli;MA Yurong;SUN Wen;WANG Lingxia;QIN Ken;LIANG Wenyu(School of Life Sciences,Ningxia University,Yinchuan 750021;State Key Laboratory of Efficient Production of Forest Resources,Yinchuan 750021;NingXia Academy of Agriculture and Forestry Sciences,Yinchuan 750002)

机构地区：[1]宁夏大学生命科学学院,银川750021 [2]林木资源高效生产全国重点实验室,银川750021 [3]宁夏农林科学院,银川750002

出　　处：《植物遗传资源学报》2025年第2期380-393,共14页Journal of Plant Genetic Resources

基　　金：2022年度宁夏留学回国人员创新创业项目;国家自然科学基金项目(32301632);宁夏自然科学基金项目(2023AAC05024)。

摘　　要：宁夏枸杞为著名耐盐药用植物,本研究以宁夏枸杞为材料,采用RACE方法克隆LbALDH3F1基因并进行序列分析,PCR法克隆LbALDH3F1基因启动子序列,构建过表达载体pCAMBIA 2300 EGFP-LbALDH3F1进行拟南芥遗传转化和亚细胞定位,并进行转LbALDH3F1拟南芥耐NaCl胁迫检验及PCR鉴定。结果表明,宁夏枸杞LbALDH3F1基因全长1700 bp,编码序列为1446 bp,编码481个氨基酸残基;上游启动子序列为1850 bp;LbALDH3F1二级结构以α螺旋和无规则卷曲为主,定位于细胞核和细胞膜中。随着NaCl胁迫程度加深,宁夏枸杞的LbALDH3F1表达量呈先升后降的趋势,且在200 mmol/L NaCl胁迫时表达量最高。对LbALDH3F1进行遗传转化,发现转基因拟南芥对NaCl胁迫的耐受能力明显增强,超氧化物歧化酶表现出更高的活性,脯氨酸及叶绿素a含量在300 mmol/L NaCl胁迫下显著升高,而过氧化氢及丙二醛等含量在高盐胁迫下显著低于野生型拟南芥,表明LbALDH3F1具有提高植物抗氧化能力从而增强耐NaCl胁迫的能力。该研究结果为深入探讨宁夏枸杞LbALDH3F1基因功能及其在响应盐胁迫过程中的应答机制奠定了基础。Lycium barbarum Linn.is a well-known salt-tolerant medicinal plant.In this study,the LbALDH3F1 gene was cloned from L.barbarum by RACE,LbALDH3F1 gene promoter sequence was also cloned by PCR.Overexpression vector pCAMBIA 2300 EGFP-LbALDH3F1 was constructed for genetic transformation and subcellular localization in Arabidopsis thaliana,detection of NaCl stress tolerance in transgenic A.thaliana and PCR identification were carried out.The results showed that the LbALDH3F1 was 1700 bp in length,with a CDS of 1446 bp,encoding 481 amino acids.The upstream promoter sequence of LbALDH3F1 was 1850 bp.The secondary structure of LbALDH3F1 was dominated byα-helix and irregular coil.The LbALDH3F1 localized in nucleus and cell membrane.With the severity of NaCl stress,the expression of LbALDH3F1 in L.barbarum showed a tendency of increasing and then decreasing,and the highest expression was found at 200 mmol/L NaCl stress.Genetic transformation of LbALDH3F1 revealed that transgenic LbALDH3F1 A.thaliana was significantly more tolerant to NaCl stress,SOD showed higher activity,proline and chlorophyll a content were significantly increased under 300 mmol/L NaCl stress,whereas H_(2)O_(2)and MDA content were significantly lower than wild type A.thaliana under high salt stress.These results lay a foundation for further studying the function of LbALDH3F1 gene in L.barbarum and its response mechanism in response to salt stress.

关 键 词：LbALDH3F1 克隆 遗传转化 盐胁迫 基因功能 宁夏枸杞 

分 类 号：S567.19[农业科学—中草药栽培]