异源过表达高粱SbSnRK2.1增强拟南芥对盐胁迫的抗性

doi:10.13560/j.cnki.biotech.bull.1985.2025-0114

摘要/Abstract

摘要：

目的从高粱基因组中发掘调控植物响应盐胁迫的SNF1 RELATED KINASE 2（SnRK2s）基因，为高粱抗盐机制研究提供理论依据。方法运用生物信息学技术对高粱SnRK2基因家族成员（SbSnRK2s）进行鉴定和序列分析，利用定量PCR技术分析SbSnRK2s在盐胁迫处理12 h内表达量的动态变化，将盐胁迫诱导显著高表达的基因在拟南芥中异源过表达，检测转基因拟南芥对盐胁迫抗性的变化。结果在高粱基因组中共鉴定到11个SbSnRK2s基因，不均匀地分布在7条染色体上，蛋白序列进化树结果表明，SbSnRK2s与同为C₄植物的玉米SnRK2亲缘关系较近；启动子分析发现，SbSnRK2s基因启动子含有激素响应、应激响应等元件。盐胁迫处理后，11个SbSnRK2s均上调表达，其中，SbSnRK2.1的上调幅度最大，在盐胁迫处理9 h后上调幅度超过150倍，在拟南芥中异源过表达SbSnRK2.1，经盐胁迫处理，转基因植株的存活率、主根长度和鲜重均显著高于野生型。结论 SbSnRK2.1在盐处理下的表达量显著升高，异源过表达SbSnRK2.1显著提高了拟南芥对盐胁迫的抗性，表明该基因在植物响应盐胁迫中具有重要作用。

关键词: SnRK2, 高粱, 盐胁迫, 异源过表达, 拟南芥

Abstract:

Objective This study aims to identify SNF1-RELATED KINASE 2 (SnRK2) genes from the sorghum genome that regulate plant responses to salt stress, thereby providing a theoretical basis for investigating salt tolerance mechanisms in sorghum. Method Bioinformatics techniques were employed to identify and analyze the sequence characteristics of sorghum SnRK2 gene family members (SbSnRK2s). Quantitative PCR was used to examine the dynamic expression patterns of SbSnRK2s within 12 h of salt stress treatment. Genes showing significant upregulation under salt stress were heterologously overexpressed in Arabidopsis thaliana to evaluate changes in salt stress tolerance. Result Eleven SbSnRK2s genes were identified in the sorghum genome, unevenly distributed across seven chromosomes. Phylogenetic analysis of protein sequences revealed close evolutionary relationships between SbSnRK2s and SnRK2 genes from maize, another C₄ plant. Promoter analysis identified regulatory elements responsive to hormones and stress. All 11 SbSnRK2s exhibited upregulated expression under salt stress, with SbSnRK2.1 showing the most pronounced induction (>150-fold increase at 9 h post-treatment). Arabidopsis thaliana heterologously expressing SbSnRK2.1 demonstrated significantly higher survival rates, primary root lengths, and fresh weights compared to wild-type plants under salt stress conditions. Conclusion SbSnRK2.1 demonstrates remarkable upregulation in response to salt treatment, and its heterologous overexpression significantly enhances the tolerance of Arabidopsis thaliana to salt stress, indicating its crucial role in plant responses to salinity stress.

Key words: SnRK2, sorghum, salt resistance, heterologous overexpression, Arabidopsis thaliana

李雅琼, 格桑拉毛, 陈启迪, 杨宇环, 何花转, 赵耀飞. 异源过表达高粱SbSnRK2.1增强拟南芥对盐胁迫的抗性[J]. 生物技术通报, 2025, 41(8): 115-123.

LI Ya-qiong, GESANG La-mao, CHEN Qi-di, YANG Yu-huan, HE Hua-zhuan, ZHAO Yao-fei. Heterologous Overexpression of Sorghum SbSnRK2.1 Enhances the Resistance to Salt Stress in Arabidopsis[J]. Biotechnology Bulletin, 2025, 41(8): 115-123.

图/表 10

表1 本研究所用的引物序列

Table 1 Primers used in this study

引物名称 Primer name	序列 Sequence （5′‒3′）
SbSnRK2.1-F	GCCTTCTTACTCAGACCAAT
SbSnRK2.1-R	TCCTTCTCTTCTTCTTCATCC
SbSnRK2.2-F	CGGCTATTCCAAGTCATCT
SbSnRK2.2-R	GCACCAACAACCATTACATA
SbSnRK2.3-F	TCATCAGTTCTACACTCTCAG
SbSnRK2.3-R	ACCGACCAGCATCACATA
SbSnRK2.4-F	AGGACGAGGCAAGGTATT
SbSnRK2.4-R	CAGCAGCGATGACTTAGA
SbSnRK2.5-F	TCAGGCTATGGCTACAGT
SbSnRK2.5-R	CACACTCATCTTCTTCTTCC
SbSnRK2.6-F	CACTCTGCTGGATGGAAG
SbSnRK2.6-R	CAATCTTGCCGTCGTATTC
SbSnRK2.7-F	TTCGCCGTCAAGTTCATC
SbSnRK2.7-R	GCCAGCAGCATATTCCATA
SbSnRK2.8-F	GTGAGAATGAGGCAAGGTT
SbSnRK2.8-F	TGAGAGTGAAGAACAGAAGAC
SbSnRK2.9-R	CATGAGGATCGTTCAGGAG
SbSnRK2.9-F	TGTCGTAGTCGTCTTCTTC
SbSnRK2.10-R	ACTTACTCCGTCCAGTCC
SbSnRK2.10-F	CCATCCTCTTCCTCCTCAT
SbSnRK2.11-R	GGCAAGAAGATTGATGAGAA
SbSnRK2.11-R	TCCACCAGCAGCATATTC

表2 高粱SnRK2基因（SbSnRK2s）基本信息

Table 2 Information of SnRK2 genes in sorghum (SbSnRK2s)

基因 Gene	基因ID Gene ID	基因长度 Gene length （bp）	CDS长度 CDS length （bp）
SbSnRK2.1	Sobic.003G370100	5 130	1 086
SbSnRK2.2	Sobic.001G078800	7 044	1 101
SbSnRK2.3	Sobic.001G168400	3 201	1 089
SbSnRK2.4	Sobic.004G173500	5 029	1 095
SbSnRK2.5	Sobic.009G149900	4 945	1 080
SbSnRK2.6	Sobic.008G147000	2 691	1 095
SbSnRK2.7	Sobic.002G379400	4 989	1 020
SbSnRK2.8	Sobic.001G350700	4 525	1 035
SbSnRK2.9	Sobic.006G279100	6 401	1 140
SbSnRK2.10	Sobic.006G083000	5 786	1 134
SbSnRK2.11	Sobic.001G294400	3 307	1 002

图1 SbSnRK2s在高粱基因组中的分布染色体上位置距离用Mb表示

Fig. 1 Distribution of SbSnRK2s in the genome of sorghumChromosomal distances are given in Mb

图2 SbSnRK2s的基因结构分析黄色部分表示外显子，黑色实线表示内含子，蓝色部分为基因上下游非编码区序列

Fig. 2 Gene structure analysis of SbSnRK2sThe yellow part indicates the exon, the black solid line indicates the intron, and the blue part indicates the sequence of the upstream and downstream non-coding regions of the SbSnRK2 genes

图3 SbSnRK2基因启动子特征分析

Fig. 3 Promoter characteristics analysis of SbSnRK2 genes

图4 SnRK2s的进化树分析Sb：高粱；At：拟南芥；Os：水稻；Zm：玉米

Fig. 4 Phylogenetic analysis of SnRK2Sb: Sorghum bicolor; At: Arabidopsis thaliana; Os: Oryza sativa; Zm: Zea mays

图5 盐胁迫下SbSnRK2s的表达模式不同小写字母表示统计学显著差异（P<0.05）

Fig. 5 Expression patterns of the SbSnRK2s under salt stressDifferent letters above the bar chart indicate significant differences at P<0.05

图6 盐胁迫下异源过表达SbSnRK2.1植株的表型（A）和存活率（B）* P<0.05. ** P<0.01. ns: No significant. Bar = 1 cm. The same below

Fig. 6 Phenotype (A) and the survival rates (B) of plants overexpressing SbSnRK2.1 under salt stress

图7 异源过表达SbSnRK2.1促进了拟南芥在盐胁迫下的主根生长A：正常生长和盐胁迫下的主根生长情况，标尺=1 cm；B：主根长度统计；C：相对根长（盐胁迫下主根长与正常生长平均根长的比值）

Fig. 7 Heterologous overexpression of SbSnRK2.1 promotes the primary root growth of A. thaliana under salt stressA: Growth of primary roots under normal and salt stress conditions, bar = 1 cm. B: Statistics of primary root length. C: Relative root length (ratio of root length under salt stress to average root length under normal growth conditions)

图8 异源过表达SbSnRK2.1和野生型拟南芥在盐胁迫下的鲜重变化

Fig. 8 Variation in fresh weights of heterologously-overexpressed SbSnRK2.1 and wild A. thaliana under salt stress

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