异源表达甜菜BvHSP18.2基因增强拟南芥对镉胁迫的耐受性研究

doi:10.11924/j.issn.1000-6850.casb2024-0265

中国农学通报 ›› 2024, Vol. 40 ›› Issue (29): 14-20.doi: 10.11924/j.issn.1000-6850.casb2024-0265

异源表达甜菜BvHSP18.2基因增强拟南芥对镉胁迫的耐受性研究

仲维婷¹^,²(), 张琼¹^,², 兴旺¹^,², 刘大丽¹^,²()

¹ 国家甜菜种质中期库/黑龙江大学，哈尔滨 150080
² 黑龙江省普通高等学校甜菜遗传育种重点实验室/黑龙江大学现代农业与生态环境学院，哈尔滨 150080

收稿日期:2024-04-23 修回日期:2024-06-15 出版日期:2024-10-15 发布日期:2024-10-14
通讯作者:
刘大丽，女，1981年出生，副研究员，研究方向：作物种质资源与抗性生理。通信地址：150080 黑龙江省哈尔滨市南岗区学府路74号黑龙江大学，Tel：0451-86608346，E-mail：daliliu_hlju@163.com。
作者简介:
仲维婷，女，2003年出生，在读研究生，研究方向：甜菜种质资源。通信地址：150080 黑龙江省哈尔滨市南岗区学府路74号黑龙江大学，Tel：0451-86608346，E-mail：15184641700@163.com。
基金资助:
财政部和农业农村部“国家现代农业产业技术体系（糖料）建设”项目(CARS-170102); 甜菜种质资源安全保存项目(19230841); 内蒙古自治区“揭榜挂帅”项目“甜菜优异种质创制与适宜机械化作业品种选育”(2022JBGS0029); 黑龙江省高校科研业务费项目(2022-KYYWF-1070); 黑龙江省自然科学基金(LH2023C090)

Heterologous Expression of BvHSP18.2 Gene in Sugar Beet Enhancing Tolerance to Cadmium of Arabidopsis thaliana

ZHONG Weiting¹^,²(), ZHANG Qiong¹^,², XING Wang¹^,², LIU Dali¹^,²()

¹ National Beet Medium-term Gene Bank/Heilongjiang University, Harbin 150080
² Key Laboratory of Sugar Beet Genetics and Breeding/College of Advanced Agriculture and Ecological Environment, Heilongjiang University, Harbin 150080

Received:2024-04-23 Revised:2024-06-15 Published:2024-10-15 Online:2024-10-14

摘要/Abstract

摘要：

为进一步分析和验证甜菜BvHSP18.2基因(LOC104903994)受镉胁迫调控的功能，本研究用RT-PCR技术克隆了甜菜BvHSP18.2基因，并通过植物表达载体构建以及拟南芥的遗传转化，测得异源表达目的基因的拟南芥植株在不同浓度镉胁迫下的表型及生理变化数据。研究结果表明，随着镉胁迫浓度的增加，异源表达BvHSP18.2基因的拟南芥无论在根长还是鲜重方面均具有优势：50 µmol/L镉胁迫下平均根长相差最大，为1.1 cm，600 µmol/L镉胁迫下平均鲜重相差最大，为0.015 g。同时，BvHSP18.2基因的过表达可有效提高拟南芥体内SOD和POD活性，300 µmol/L镉胁迫下SOD活性最高，为657.7266953 U/g；600 µmol/L镉胁迫下POD活性最高，为野生型拟南芥的1.91倍。BvHSP18.2基因的过表达增强了拟南芥在镉胁迫下的耐受性，推测该基因在植物对镉胁迫的适应机制中具有重要的作用。

关键词: 甜菜, 拟南芥, BvHSP18.2基因, 异源表达, 镉胁迫, 功能分析, 基因克隆, SOD活性, POD活性, 耐受性, 遗传转化

Abstract:

In order to further analyze and verify the function of the sugar beet BvHSP18.2 gene (LOC104903994) regulated by cadmium stress, we cloned the sugar beet BvHSP18.2 gene by RT-PCR in this study, and measured the phenotypic and physiological phenotypes and physiological changes of Arabidopsis thaliana plants heterologously expressing the target gene under different concentrations of cadmium stress by plant expression vector construction as well as Arabidopsis thaliana genetic transformation. The phenotypic and physiological changes of Arabidopsis thaliana plants heterologously expressing the target gene under different concentrations of Cd were measured. The results showed that with the increase of cadmium stress concentration, Arabidopsis thaliana plants heterologously expressing the BvHSP18.2 gene had an advantage in both root length and fresh weight. The largest difference was 1.1 cm in average root length under 50 µmol/L cadmium stress, and the largest difference was 0.015 g in average fresh weight under 600 µmol/L cadmium stress, and the overexpression of the BvHSP18.2 gene could effectively increase the S-phase of Arabidopsis thaliana plants under different concentrations of cadmium stress. The overexpression of BvHSP18.2 gene enhanced the tolerance of Arabidopsis thaliana under cadmium stress, and it was hypothesized that this gene played an important role in the adaptation of plants to cadmium stress.

Key words: sugar beet, Arabidopsis thaliana, BvHSP18.2 gene, heterologous expression, cadmium stress, functional analysis, gene cloning, sod activity, pod activity, tolerance, genetic transformation

仲维婷, 张琼, 兴旺, 刘大丽. 异源表达甜菜BvHSP18.2基因增强拟南芥对镉胁迫的耐受性研究[J]. 中国农学通报, 2024, 40(29): 14-20.

ZHONG Weiting, ZHANG Qiong, XING Wang, LIU Dali. Heterologous Expression of BvHSP18.2 Gene in Sugar Beet Enhancing Tolerance to Cadmium of Arabidopsis thaliana [J]. Chinese Agricultural Science Bulletin, 2024, 40(29): 14-20.

图/表 6

图1. 甜菜BvHSP18.2基因的克隆 (A)：甜菜总RNA；(B)：M (DL 5000)，RT-PCR扩增BvHSP18.2；(C)：M (DL 1000)；1~2：pEASY-Blunt-BvHSP18.2 PCR鉴定结果；(D)：M (DL 5000)，pEASY-Blunt-BvHSP18.2 BamHI及XhoI双酶切鉴定

图2. pCAMBIA1300-BvHSP18.2-GFP载体构建 (A)：M (DL5000)，1：pCAMBIA1300-BvHSP18.2-GFP PCR鉴定；(B)：M (DL 5000)；1：pCAMBIA1300-GFP的PstI和SalI双酶切

图3. 转基因拟南芥验证 M：DL5000；1：野生型拟南芥 2：BvHSP18.2-OS；3：重组质粒pCAMBIA1300-BvHSP18.2-GFP

图4. 不同浓度镉胁迫下，过表达BvHSP18.2拟南芥的生长情况

图5. 不同浓度镉胁迫下，过表达BvHSP18.2拟南芥的根长及鲜重变化数据为3次独立试验的“平均值±SD”，不同字母表示差异显著(P<0.05)

图6. 不同浓度镉胁迫下，过表达BvHSP18.2拟南芥的SOD及POD酶活性数据为3次独立试验的“平均值±SD”，不同字母表示差异显著(P<0.05)

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异源表达甜菜BvHSP18.2基因增强拟南芥对镉胁迫的耐受性研究

Heterologous Expression of BvHSP18.2 Gene in Sugar Beet Enhancing Tolerance to Cadmium of Arabidopsis thaliana

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