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Acta Agronomica Sinica ›› 2025, Vol. 51 ›› Issue (7): 1736-1746.doi: 10.3724/SP.J.1006.2025.44141

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Cloning and functional analysis of glutathione S-transferase gene IbGSTU7 in sweetpotato

YIN Yu-Meng1**(), WANG Yan-Nan1**(), KANG Zhi-He1, QIAO Shou-Chen1, BIAN Qian-Qian1, LI Ya-Wei1, CAO Guo-Zheng1, ZHAO Guo-Rui1, XU Dan-Dan2, YANG Yu-Feng1,*()   

  1. 1Cereal Crop Research Institute, Henan Academy of Agricultural Sciences, Zhengzhou 450002, Henan, China
    2Tongxu Academy of Agricultural Sciences, Kaifeng 475400, Henan, China
  • Received:2024-08-30 Accepted:2025-04-27 Online:2025-07-12 Published:2025-05-07
  • Contact: *E-mail: yyfyyf5@163.com
  • About author:**Contributed equally to this work
  • Supported by:
    China Agriculture Research System of MOF and MARA(CARS-10);Special Fund for Henan Agriculture Research System(HARS-22-04-G1);Outstanding Youth Science and Technology Fund of Henan Academy of Agricultural Sciences(2024YQ17);Autonomous Innovation Project of Henan Academy of Agricultural Sciences(2024ZC014);Central Government Guides Local Funds for Scientific and Technological Development of Henan Province(Z20231811177)

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Abstract:

In this study, the IbGSTU7 gene was cloned based on our previously obtained transcriptome data from a sweetpotato skin color mutant. The open reading frame (ORF) of IbGSTU7 is 660 bp in length and encodes a protein comprising 219 amino acids. Protein sequence analysis revealed that IbGSTU7 is an acidic, hydrophilic, and structurally stable protein, showing the closest phylogenetic relationship to its homolog in Ipomoea triloba, a wild relative of sweetpotato. Subcellular localization analysis indicated that IbGSTU7 is localized in the cytoplasm. Quantitative RT-PCR analysis demonstrated that IbGSTU7 is expressed across major sweetpotato tissues, with the highest transcript level observed in the skin of storage roots. Furthermore, its expression was inducible by drought, salt, and exogenous abscisic acid (ABA) treatments. The IbGSTU7 gene was subsequently overexpressed in sweetpotato using an Agrobacterium tumefaciens-mediated transformation method. In overexpression (OE) lines, anthocyanin content in the skin of storage roots and the expression levels of anthocyanin biosynthesis-related genes IbPAL and IbUFGT were significantly increased. Additionally, the OE plants exhibited enhanced tolerance to PEG-simulated drought stress, as evidenced by significant increases in fresh weight and root length compared to wild-type (WT) plants. Under drought conditions, OE lines showed a marked reduction in hydrogen peroxide (H2O2) levels, accompanied by significant upregulation of reactive oxygen species (ROS) scavenging-related genes IbMDHAR and IbPOD. These findings provide novel insights into the role of IbGSTU7 in regulating anthocyanin accumulation and improving drought stress tolerance in sweetpotato.

Key words: sweetpotato, IbGSTU7, overexpression, anthocyanin, drought resistance

Table 1

Primers used in this study"

引物名称
Primer name		 引物序列
Primer sequence (5′-3′)		 引物目的
Primer purpose
IbORF-F ATGGCAGAAGTGAAGGTTTT ORF扩增引物
IbORF-R TTACATGAATACCTTGGTGTAGTAG ORF amplification
GSTU7-p1300-F CGAGCTCATGGCAGAAGTGAAGGTTTTAG 亚细胞定位
GSTU7-p1300-R CGGATCCCATGAATACCTTGGTGTAGTAGGG Subcellular localization
GSTU7-p1302-F GACTAGTATGGCAGAAGTGAAGGTTTT 过表达
GSTU7-p1302-R GGGTGACCTTACATGAATACCTTGGTGTAGTAG Overexpression
1302-35S-F GAAGATAGTGGAAAAGGAAGGTG 转基因阳性鉴定
GSTU7-324-R CCTTTGGTAAATACTGCTTTCTCC Transgenic positive identification
QGSTU7-F AGTGAAGGTTTTAGGGTCGTGG 实时荧光定量PCR
QGSTU7-R GACAGGGTTGGATTTGAGAAGC Real-time quantitative PCR
Actin-F AGCAGCATGAAGATTAAGGTTGTAGCAC
Actin-R TGGAAAATTAGAAGCACTTCCTGTGAAC
IbMDHAR-F CTACTCCCGTGCCTTTGATT
IbMDHAR-R CTCCAAGAATGCACCAACAA
IbPOD-F TTCACGACTGCTTCGTTGA
IbPOD-R TTCTCAACCGCGGTCTTAA
IbCHI-F CCGATCATAGAAGCCGCCAT
IbCHI-R CTCCGGCATTGAACCCTCTT
IbPAL-F CCCTGCAGTGCTAACTACCC
IbPAL-R GAATAGCCGGGTTCCCACTC
IbUFGT-F CAAACGGAAACGGGTTGGAC
IbUFGT-R CGGTGGGTTCTAGCTTCTGG

Fig. 1

Bioinformatic analysis of the IbGSTU7 gene A: tertiary structure analysis of the IbGSTU7 protein; B: phylogenetic analysis of IbGSTU7 and homologous proteins from other species."

Fig. 2

Subcellular localization of the IbGSTU7 protein Scale: 30.0 μm."

Fig. 3

Expression analysis of the IbGSTU7 gene in Sushu 8 A: expression analysis of IbGSTU7 gene in different tissues of Sushu 8. B: the response of IbGSTU7 to different treatments. S: stems; L: leaves; PR: pencil roots; SSR: the skin of storage roots; FSR: the flesh of storage roots. *, ** indicate significant differences at the 0.05 and 0.01 probability levels, respectively."

Fig. 4

Production of transgenic sweetpotato plants overexpressing the IbGSTU7 gene A: embryogenic calluses of sweetpotato cv. Lizixiang; B: formation of Hyg-resistant callus; C: formation of somatic embryos formation; D: regeneration of putatively transgenic plants; E: enlarged transgenic plants; F: whole putatively transgenic plants; G: propagation of test-tube seedlings; H: greenhouse domestication; I: field planting. Scale: 2 cm."

Fig. 5

Expression analysis of IbGSTU7 gene in OE transgenic sweetpotato plants by qRT-PCR *, ** indicate significant differences at the 0.05 and 0.01 probability levels, respectively."

Fig. 6

Overexpression of IbGSTU7 gene promotes the total anthocyanin accumulation in the skin of storage roots A: skin and flesh color of WT and IbGSTU7-OE plants after four months’ growth in the field, Scale: 5 cm; B-C: contents of carotenoids (B) and total anthocyanins (C) in the skin of storage roots of WT and IbGSTU7-OE plants; D: contents of total anthocyanins in the flesh of storage roots of WT and IbGSTU7-OE plants; E-G: expression levels of IbCHI (E), IbPAL (F), and IbUFGT (G) genes of WT and IbGSTU7-OE plants. *, ** indicate significant differences at the 0.05 and 0.01 probability levels, respectively."

Fig. 7

Overexpression of IbGSTU7 gene enhances drought tolerance in sweetpotato A-F: phenotypes (A), fresh weight (B), root length (C), IbMDHAR gene expression level (D), IbPOD gene expression level (E) and H2O2 content (F) of WT and IbGSTU7-OE plants cultivated on MS Medium with 20% PEG-6000 treatment for four weeks. Scale: 5 cm. *, ** indicate significant differences at the 0.05 and 0.01 probability levels, respectively."

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[5] Wang Yongsheng;Wang Jing;Duan Jingya;Wang Jinfa;Liu Liangshi. Isolation and Genetic Research of a Dwarf Tiilering Mutant Rice[J]. Acta Agron Sin, 2002, 28(02): 235 -239 .
[6] WANG Li-Yan;ZHAO Ke-Fu. Some Physiological Response of Zea mays under Salt-stress[J]. Acta Agron Sin, 2005, 31(02): 264 -268 .
[7] TIAN Meng-Liang;HUNAG Yu-Bi;TAN Gong-Xie;LIU Yong-Jian;RONG Ting-Zhao. Sequence Polymorphism of waxy Genes in Landraces of Waxy Maize from Southwest China[J]. Acta Agron Sin, 2008, 34(05): 729 -736 .
[8] HU Xi-Yuan;LI Jian-Ping;SONG Xi-Fang. Efficiency of Spatial Statistical Analysis in Superior Genotype Selection of Plant Breeding[J]. Acta Agron Sin, 2008, 34(03): 412 -417 .
[9] WANG Yan;QIU Li-Ming;XIE Wen-Juan;HUANG Wei;YE Feng;ZHANG Fu-Chun;MA Ji. Cold Tolerance of Transgenic Tobacco Carrying Gene Encoding Insect Antifreeze Protein[J]. Acta Agron Sin, 2008, 34(03): 397 -402 .
[10] ZHENG Xi;WU Jian-Guo;LOU Xiang-Yang;XU Hai-Ming;SHI Chun-Hai. Mapping and Analysis of QTLs on Maternal and Endosperm Genomes for Histidine and Arginine in Rice (Oryza sativa L.) across Environments[J]. Acta Agron Sin, 2008, 34(03): 369 -375 .
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