花生抗旱基因AhCPK8及其互作蛋白GAPDH的克隆及分子鉴定

doi:10.3724/SP.J.1006.2025.55028

Abstract

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

HE Mei, ZHANG Jia-Lei, FAN Shi-Kai, MENG Jing-Jing, WANG Jian-Guo, GUO Feng, LI Xin-Guo, WAN Shu-Bo, YANG Sha. Cloning and molecular identification of the peanut drought-resistance gene AhCPK8 and its interacting protein GAPDH[J].Acta Agronomica Sinica, 2025, 51(10): 2713-2726.

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基因名称 Gene name	正向引物序列 Forword primer sequence (5′-3′)	反向引物序列 Reverse primer sequence (5′-3′)
TUA5	CTGATGTCGCTGTGCTCTTGG	CTGTTGAGGTTGGTGTAGGTAGG
AhCPK2	TGGGCAGAGACCGAACAAGGG	CCGCTGTCAACCGCTTCTTAGG
AhCPK8	CCTGCAAGTCCATCCCAAAGCG	CGAGATGAACGGCAAGGCTGTC
AhCPK36	TTCCTCCTCATCCGCCACCATC	CCCAGCTCCCTACCGAAGACG
AhCPK40	TGGAGGCGGCTGATGTGGAC	CCTCGCGCTCTAGCTTGTTGAG

名称 Symbol	序列ID Sequence ID	名称 Symbol	序列ID Sequence ID
AhGAPA1	arahy.Tifrunner.gnm2.ann1.D2DMXG.1	AhGAPC3	arahy.Tifrunner.gnm2.ann1.8UAX1T.1
AhGAPA2	arahy.Tifrunner.gnm2.ann1.YN8IJ2.1	AhGAPC4	arahy.Tifrunner.gnm2.ann1.YH7XEC.1
AhGAPA3	arahy.Tifrunner.gnm2.ann1.8EQN3M.1	AhGAPC5	arahy.Tifrunner.gnm2.ann1.YH7XEC.3
AhGAPA4	arahy.Tifrunner.gnm2.ann1.HFT5ZK.1	AhGAPC6	arahy.Tifrunner.gnm2.ann1.SIHD8Y.1
AhGAPA5	arahy.Tifrunner.gnm2.ann1.FN6RIW.1	AhGAPC7	arahy.Tifrunner.gnm2.ann1.VRBE94.1
AhGAPA6	arahy.Tifrunner.gnm2.ann1.BA5EUQ.1	AhGAPC8	arahy.Tifrunner.gnm2.ann1.QHP0ED.1
AhGAPA7	arahy.Tifrunner.gnm2.ann1.BFW5UL.1	AhGAPCp1	arahy.Tifrunner.gnm2.ann1.5PGU74.1
AhGAPA8	arahy.Tifrunner.gnm2.ann1.6W2CKZ.1	AhGAPCp2	arahy.Tifrunner.gnm2.ann1.5PGU74.2
AhGAPA9	arahy.Tifrunner.gnm2.ann1.ZX8DBB.1	AhGAPCp3	arahy.Tifrunner.gnm2.ann1.F06IQB.2
AhGAPC1	arahy.Tifrunner.gnm2.ann1.YL3Z6V.1	AhGAPCp4	arahy.Tifrunner.gnm2.ann1.F06IQB.3
AhGAPC2	arahy.Tifrunner.gnm2.ann1.PV44JT.1

名称 Symbol	氨基酸数目 Number of amino acid	分子量 Molecular weight (Da)	等电点 pI	不稳定指数 Instability index	脂溶指数 Aliphatic index	总平均亲水性值 Grand average of hydropathicity
AhGAPA1	388	41,339.06	8.92	24.19	89.95	-0.079
AhGAPA2	509	54,458.07	5.59	29.04	89.80	-0.101
AhGAPA3	359	38,725.28	5.48	24.14	93.31	-0.040
AhGAPA4	428	45,980.96	9.40	41.61	92.69	-0.130
AhGAPA5	405	43,179.17	8.44	24.73	91.68	-0.056
AhGAPA6	478	51,430.76	6.80	27.71	89.08	-0.114
AhGAPA7	381	40,951.94	5.50	25.43	94.88	-0.005
AhGAPA8	469	50,071.35	5.32	32.03	99.10	0.082
AhGAPA9	586	63,306.33	7.93	41.38	91.96	-0.102
AhGAPC1	336	36,546.89	7.06	23.08	91.34	-0.108
AhGAPC2	347	37,480.97	8.23	33.58	90.49	-0.065
AhGAPC3	278	31,231.71	6.07	30.71	90.43	-0.282
AhGAPC4	339	36,841.07	7.02	28.17	90.27	-0.087
AhGAPC5	223	24,220.73	8.68	26.63	83.50	-0.122
AhGAPC6	336	36,250.48	8.24	31.23	89.67	-0.080
AhGAPC7	336	36,532.86	7.06	23.33	91.04	-0.109
AhGAPC8	339	36,855.10	7.02	27.16	90.27	-0.087
AhGAPCp1	416	44,435.84	9.00	32.74	89.30	-0.039
AhGAPCp2	345	37,141.40	6.61	29.29	86.75	-0.051
AhGAPCp3	440	47,250.28	9.06	33.15	89.52	-0.027
AhGAPCp4	416	44,479.96	9.04	31.08	89.06	-0.044

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

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