Function Analysis ofStRGLG1in Regulating Potato to Drought Stress Response

doi:10.16420/j.issn.0513-353x.2024-0960

Acta Horticulturae Sinica ›› 2025, Vol. 52 ›› Issue (9): 2329-2342.doi: 10.16420/j.issn.0513-353x.2024-0960

• Genetic & Breeding·Germplasm Resources·Molecular Biology • Previous Articles Next Articles

Function Analysis ofStRGLG1in Regulating Potato to Drought Stress Response

ZHU Cunlan¹^,², FAN Junliang¹^,², WANG Kaitong¹^,³, WEI Meng¹^,², YANG Liang¹^,³, LIU Haotian¹^,³, SI Huaijun¹^,², ZHANG Ning¹^,²^,^*()

¹ State Key Laboratory of Aridland Crop Science， Gansu Agricultural University， Lanzhou 730070， China
² College of Life Science and Technology， Gansu Agricultural University， Lanzhou 730070， China
³ College of Agronomy， Gansu Agricultural University， Lanzhou 730070， China

Received:2025-04-18 Revised:2025-07-21 Online:2025-09-25 Published:2025-09-24
Contact: ZHANG Ning

Abstract

Abstract:

RING DOMAIN LIGASE（RGLG）is a ring-type ubiquitin ligase E3，which is involved in plant growth and development，signal transduction and abiotic stress. To further investigate the function and regulatory mechanism ofStRGLG1gene in potato stress response，qRT-PCR was used to assay the expression level ofStRGLG1gene in potato under drought stress.StRGLG1overexpression and interference expression plants of potato were obtained by genetic transformation. The activities of superoxide dismutase（SOD），catalase（CAT）and peroxidase（POD）and the content of malondialdehyde（MDA）in transgenic plants were measured before and after drought stress. StRGLG1 interacting proteins were identified and validated by yeast two-hybrid（Y2H）and bimolecular fluorescence complementation（BiFC）. The results showed that SOD，POD and CAT activities ofStRGLG1overexpression plants increased by 31.78%，46.87% and 51.76% on average，and MDA content decreased by 23.12%. SOD，POD and CAT decreased by 15.83%，18.28% and 13.78% respectively，while MDA content increased by 13.30% under drought stress. It was proved thatStRGLG1gene played a role in the regulation of potato drought tolerance. Y2H and BiFC results showed that StRGLG1 and StERF53 proteins interact. These findings indicate thatStRGLG1gene expression plays a role in potato stress resistance，and the results can provide references for the molecular mechanism ofStRGLG1regulation of potato drought stress response.

Key words: potato, StRGLG1, drought stress, protein interaction

ZHU Cunlan, FAN Junliang, WANG Kaitong, WEI Meng, YANG Liang, LIU Haotian, SI Huaijun, ZHANG Ning. Function Analysis ofStRGLG1in Regulating Potato to Drought Stress Response[J]. Acta Horticulturae Sinica, 2025, 52(9): 2329-2342.

Figures/Tables 7

Table 1 Primers used in this study

基因名称/ID Gene name/ID	引物序列（5′-3′） Primer sequence		用途 Application
pCAMBIA1300‐3 5S‐StRGLG1	F	AGAACACGGGGGACGAGCTCATGGGGAATCAAGAGTCTGC	载体构建 Construction of vector
pCAMBIA1300‐3 5S‐StRGLG1	R	TGCTCACCATGTCGACCTAGCTATATAGTCTTATGCGAGTT
pEGFP-amiR-StRGLG1	I miR-s	gaTATGGAATTTACAGTAGCCTAtctctcttttgtattcc
	II miR-a	gaTAGGCTACTGTAAATTCCATAtcaaagagaatcaatga
	III miR*s	gaTAAGCTACTGTAATTTCCATTtcacaggtcgtgatatg
	IV miR*a	gaAATGGAAATTACAGTAGCTTAtctacatatatattcct
	A	CTGCAAGGCGATTAAGTTGGGTAAC
	B	CTGTTTCCTGTGTGAAATTGTTATCCGC
pGBKT7‐StRGLG1	F	AGGAGGACCTGCATATGATGGGGAATCAAGAGTCTGC
pGBKT7‐StRGLG1	R	GCCGCTGCAGGTCGACCTAGCTATATAGTCTTATGCGAGTT
pGADT7‐StERF53	F	GGAGGCCAGTGAATTATGGCAGCTATGGATTTTTGGAA
pGADT7‐StERF53	R	TTCATCTGCAGCTCGTTATAGGGAGGCCCAATCAATTTCA
pSPYNE-StRGLG1	F	CACGGGGGACTCTAGATGGGGAATCAAGAGTCTGC
pSPYNE-StRGLG1	R	TCCATCCCGGGAGCGGCTATATAGTCTTATGCGAGTTTT
pSPYCE-StERF53	F	CACGGGGGACTCTAGATGGCAGCTATGGATTTTTGGAA
pSPYCE-StERF53	R	TACATCCCGGGAGCGTAGGGAGGCCCAATCAATTTCATGA
StRGLG1	F	ACCACACACGATCAGCATGT	qRT-PCR
StRGLG1	R	GTTGTGTGGCTCGGTACTGA
StERF53	F	CAGACTGCGCATCATGCTTC
StERF53	R	TGTCCCAAGTTGGCTCTTCC
Ef1α	F	GATGGTCAGACCCGTGAACA	内参基因 Reference gene
Ef1α	R	CCTTGGAGTACTTCGGGGTG	内参基因 Reference gene
HYG	F	GCTTCTGCGGGCGATTTGTGT	转基因植株鉴定 Transgenic plant identification
HYG	R	GGTCGCGGAGGCTATGGATGC	转基因植株鉴定 Transgenic plant identification

Fig. 1 The relative expression level of StRGLG1 in different potato tissues（A）and the relative expression level of StRGLG1 gene under drought stress（B） WS1：Water stress group 1（75%-85%）；WS2：Water stress group 2（55%-65%）；WS3：Water stress group 3（35%-45%）；WS4：Water stress group 4（15%-25%）. Different lowercase letters indicated significant differences （n=3；P< 0.05）. The same below

Fig. 2 Acquisition and identification of transgenic plants A，B：Calli；C，D：Differentiated buds；E，F：Rooting and screening of transgenic plants；G，H：PCR detection of transgenic plants；I，J：Relative expression levels ofStRGLG1in transgenic plants and non-transgenic plants（NT）. M：DL 2000 molecular marker；1：Positive control plasmid；2：Negative control of non-transgenic plants；3-5：Transgenic lines. A，C，E，G，I：Plant transformation process carrying recombinant plasmid pCAMBIA1300-35S-StRGLG1；B，D，F，H，J：Plant transformation process carrying recombinant plasmid pEGFP-amiR-StRGLG1

Fig. 3 Phenotype（A）and determination of physiological and biochemical indexes（B）of transgenic potato at 21 days under drought stress * and ** respectively represent the significant difference levels and extremely significant difference levels between different strains and non-transgenic plants *P< 0.05；**P< 0.01，n= 3

Fig. 4 Toxicity detection of bait vector pGBKT7-StRGLG1（A）and self-activation detection of decoy vector pGBKT7-StRGLG1（B） 1，2：Positive control pGADT7-RecT + pGBKT7-53；3，4：Negative control pGADT7-RecT + pGBKT7-Lam；5，6：Experimental group pGADT7 + pGBKT7-StRGLG1

Fig. 5 Yeast two-hybrid experiment and BiFC verification A：StRGLG1 and StERF53 one-on-one validation；B：The interaction between StRGLG1 and StERF53 in tobacco was detected by BiFC analysis

Fig. 6 The relative expression of StERF53 gene in leaves of non-transgenic plants and transgenic StRGLG1 plants under drought stress was detected by qRT-PCR n= 3；*P< 0.05；**P< 0.01

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Function Analysis ofStRGLG1in Regulating Potato to Drought Stress Response

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