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花生抗旱基因AhCPK8及其互作蛋白GAPDH的克隆及分子鉴定

何梅1,2，张佳蕾1，范士凯1，孟静静1，王建国1，郭峰1，李新国1，万书波1,*，杨莎1,*   

  1. 1 山东省农业科学院农作物种质资源研究所, 山东济南 250100; 2 山东师范大学生命科学学院, 山东济南 250399
  • 收稿日期:2025-04-17 修回日期:2025-07-09 接受日期:2025-07-09 网络出版日期:2025-07-22
  • 基金资助:
    本研究由国家自然科学基金项目(32272020), 山东省重点研发计划项目(ZFJH202310)和泰山学者工程项目(tspd20221107, tsqn202408305)资助。

Cloning and molecular identification of the peanut drought-resistance gene AhCPK8 and its interacting protein GAPDH

HE Mei1,2,ZHANG Jia-Lei1,FAN Shi-Kai1,MENG Jing-Jing1,WANG Jian-Guo1,GUO Feng1,LI Xin-Guo1,WAN Shu-Bo1,*,YANG Sha1,*   

  1. 1 Crop Germplasm Resources Institute, Shandong Academy of Agricultural Sciences, Jinan 250100, Shandong, China; 2 College of Life Sciences, Shandong Normal University, Jinan 250399, Shandong, China
  • Received:2025-04-17 Revised:2025-07-09 Accepted:2025-07-09 Published online:2025-07-22
  • Supported by:
    This study was supported by the National Natural Science Foundation of China (32272020), the Shandong Key R&D Program (ZFJH202310), and the Taishan Scholars Program (tspd20221107, tsqn202408305).

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摘要：

花生(Arachis hypogaea L.)是重要的油料和经济作物，干旱是限制花生产量的主要因素之一。探明花生响应干旱胁迫的关键基因，对后续花生品种改良，提高产量具有重要意义。本研究以花生品种花育22号(HY22)为研究对象，足钙(SC)、缺钙(NC) 2种营养液水培培养，终浓度为20% PEG-6000溶液模拟干旱胁迫处理(DSC、DNC)。通过干旱处理后转录组数据结合花生钙传感蛋白编码基因CPKs的差异表达筛选出花生抗旱基因AhCPK8。以干旱处理后的花生叶片为材料，利用酵母双杂交筛选到其互作蛋白甘油醛-3-磷酸脱氢酶(GAPDH)并得到初步验证。GAPDH家族基因在调节植物对非生物胁迫的反应中发挥着重要作用。基于AhGAPDH基因家族的生物信息学分析，在花生中共鉴定出21个GAPDH基因，再根据大豆和拟南芥系统发育树关系以及基因在12条染色体上的先后顺序进行命名。基本理化特征结果表明，AhGAPDH家族成员大多数为稳定、疏水性蛋白；同一聚类的AhGAPDH家族成员大多数表现出相似的motif结构、保守结构域和基因结构；启动子区域中有丰富的光响应、植物激素响应及非生物胁迫响应相关的顺式作用元件。AhCPK8与AhGAPDH存在相互作用，共同调控花生的抗旱性。

关键词: 花育22号, 抗旱性, AhCPK8, 互作蛋白, GAPDH

Abstract:

Peanut is an important oilseed and economic crop, and drought is one of the major factors limiting its yield. Identifying key genes involved in the peanut drought response is of great significance for improving drought tolerance and enhancing yield. In this study, the peanut variety Huayu 22(HY22) was grown hydroponically using two nutrient solutions: one with sufficient calcium (SC) and one without calcium (NC). Drought stress treatments (DSC and DNC) were simulated using PEG-6000 at a final concentration of 20%. Based on transcriptome analysis and differential expression of calcium-dependent protein kinases (CPKs), we identified the drought-responsive gene AhCPK8, which encodes a calcium-sensing protein in peanut. Using yeast two-hybrid assays and drought-treated peanut leaves, we screened for and validated its interacting protein, glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The GAPDH gene family plays a crucial role in regulating plant responses to abiotic stress. Through bioinformatics analysis, a total of 21 AhGAPDH genes were identified in peanut. These genes were named according to their phylogenetic relationships with soybean and Arabidopsis thaliana, as well as their chromosomal positions across the 12 peanut chromosomes. Physicochemical characterization revealed that most AhGAPDH proteins are stable and hydrophobic. Members within the same phylogenetic cluster shared similar motif structures, conserved domains, and gene architectures. Moreover, their promoter regions contain numerous cis-acting elements related to light responsiveness, phytohormone signaling, and abiotic stress responses. The interaction between AhCPK8 and AhGAPDH suggests a coordinated regulatory mechanism that contributes to drought resistance in peanut.

Key words: Huayu 22, drought resistance, AhCPK8, interacting protein, GAPDH

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