Scientia Agricultura Sinica ›› 2022, Vol. 55 ›› Issue (6): 1139-1148.doi: 10.3864/j.issn.0578-1752.2022.06.007

• PLANT PROTECTION • Previous Articles     Next Articles

The Role of Chalcone Synthase Gene in Grape Resistance to Gray Mold and Downy Mildew

GUO ZeXi(),SUN DaYun,QU JunJie,PAN FengYing,LIU LuLu,YIN Ling()   

  1. Key Lab of Guangxi Crop Genetic Improvement and Biotechnology, Guangxi Academy of Agricultural Sciences, Nanning 530007
  • Received:2021-09-23 Accepted:2021-12-14 Online:2022-03-16 Published:2022-03-25
  • Contact: Ling YIN E-mail:1162410925@qq.com;779335723@qq.com

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Abstract:

【Objective】The objective of this study is to clone the chalcone synthase gene CHS1 in different resistant grape varieties, clarify the expression pattern under the infection of Botrytis cinerea and Plasmopara viticola, verify the subcellular localization and function of CHS1, and to provide a basis for exploring the broad-spectrum resistance mechanism of CHS1.【Method】CHS1 genes were cloned from the leaf cDNAs of Vitis quinquangularis ‘Yeniang-2’, Vitis vinifera ‘Thompson Seedless’ and Vitis amurensis ‘Shuanghong’. The differences in the nucleic acid sequence and physicochemical properties of CHS1 protein were analyzed by bioinformatics. Expression patterns of CHS1 in different varieties during B. cinerea and P. viticola infection were also detected by qRT-PCR. The expression vector was constructed, and the transient expression technology mediated by Agrobacterium was used to verify the subcellular localization and disease resistance function in the leaves of tobacco and ‘Thompson Seedless’ tissue culture seedlings.【Result】The CHS1 is highly conserved among different grape varieties. The full length of ORF is 1 182 bp, which encodes 393 amino acids. The predicted molecular weight of the encoded protein is 42.92 kD, which is a stable hydrophilic protein. After being infected by B. cinerea, the expression patterns of VqCHS1 and VvCHS1 were similar, but the expression level of VqCHS1 in the resistant variety ‘Yejiang-2’ was significantly higher. After being infected by P. viticola, VaCHS1 and VvCHS1 showed different expression pattern changes. The expression level of VaCHS1 in the resistant variety ‘Shuanghong’ continued to increase, while the expression level of VvCHS1 gradually decreased. The results of subcellular localization showed that CHS1 protein is distributed in the cytoplasm and nucleus, but it is mainly localized in the nucleus. Compared with the negative control, the ‘Thompson Seedless’ leaves overexpressing VaCHS1 showed stronger resistance to gray mold and downy mildew after artificial inoculation. 【Conclusion】The expression level of CHS1 in resistant varieties is higher than that in susceptible varieties, and overexpression of CHS1 can increase the resistance of susceptible grape varieties to gray mold and downy mildew.

Key words: chalcone synthase (CHS), grape, Botrytis cinerea, Plasmopara viticola, disease resistance

Table 1

Primer sequences used in this study"

引物名称 Primer name 序列Primer sequence (5′-3′) 用途Purpose
CHS1-F ATGGTGTCAGTGGGGGAAAT 基因扩增
Gene amplification
CHS1-R TGCTACACAATCGACTCAC
CHS1-qF CGTTCTGAGCGAGTATGGGA 实时荧光定量PCR
Real-time fluorescence quantitative PCR
CHS1-qR TGTGGTGCCCTTTCCTTCTT
EF-1α-F AATTTTGACCAAGATCGACAGG 葡萄内参基因
Reference gene in grape
EF-1α-R CAGCAACAGTTTGACGCATG
CHS1-2F GCTCTAGAATGGTGTCAGTGGGGGAAATC 亚细胞定位
Subcellular localization
CHS1-2R GCCCCGGGATGTGAGTCGATTGTGTAGCA

Fig. 1

Phenotypes of leaf discs of different grape varieties inoculated with P. viticola and B. cinerea"

Fig. 2

Amino acid sequence alignment of VqCHS1, VvCHS1 and VaCHS1"

Fig. 3

The expression changes of CHS1 in ‘Yeniang-2’ and ‘Thompson Seedless’ during B. cinerea infection Different lowercase letters on the bars indicate significant difference in the relative expression at 0.05 level. The same as Fig. 4"

Fig. 4

The expression changes of CHS1 in ‘Shuanghong’ and ‘Thompson Seedless’ during P. viticola infection"

Fig. 5

Subcellular localization of CHS protein in tobacco leaves"

Fig. 6

Function verification of CHS1 in disease resistance"

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