Cloning，Expression，and Functional Research of BhYAB4b in Wax Gourd

doi:10.16420/j.issn.0513-353x.2024-0720

Acta Horticulturae Sinica ›› 2025, Vol. 52 ›› Issue (9): 2353-2362.doi: 10.16420/j.issn.0513-353x.2024-0720

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Cloning，Expression，and Functional Research of BhYAB4b in Wax Gourd

WANG Baochen¹^,², LUO Chen¹, YAN Jinqiang¹, CHENG Xiaoxin¹, MO Renlian¹, LIU Wenrui¹, ZHANG Yuyang², JIANG Biao¹^,^*()

¹ Guangdong Key Laboratory for New Technology Research on Vegetables/Vegetable Research Institute， Guangdong Academy of Agricultural Sciences， Guangzhou 510640， China
² National Key Laboratory for Germplasm Innovation & Utilization of Horticultural Crops， College of Horticulture & Forestry，Huazhong Agricultural University， Wuhan 430070， China

Received:2024-11-12 Revised:2025-04-24 Online:2025-09-25 Published:2025-09-24
Contact: JIANG Biao

Abstract

Abstract:

The YABBY gene family encodes a unique class of transcription factors in higher plants，which plays important regulatory roles in plant growth and development. In this study，a YABBY gene named BhYAB4b was cloned from wax gourd and this gene was subjected to bioinformatics analysis，expression pattern analysis，subcellular localization analysis，and heterologous overexpression in Arabidopsis analysis. The length of the coding sequence of BhYAB4b was 624 bp，and the predicted molecular weight of the encoded protein was 23.469 kD，containing a conserved structural domain of C2C2 zinc finger structure at the N-terminal end and a conserved structural domain of YABBY at the C-terminal end. Phylogenetic analysis showed that the BhYAB4b protein was closely related to the YAB4 proteins of watermelon and bitter melon，qRT-PCR results showed that BhYAB4b was expressed specifically in wax gourd. In the advanced-generation inbred line B227 of wax gourd，BhYAB4b was expressed in the roots，stems，leaves，male and female flowers，and fruits，while its expression in fruits was significantly higher than that in other tissues，and its expression in fruits was higher on the day of pollination than other two periods. Subcellular localization results showed that the BhYAB4b protein was localized in the nucleus of the cell. When the BhYAB4b gene was heterologous overexpression in Arabidopsis，the transgenic lines showed larger and more rosette leaves，along with enhanced fruit length and width，suggesting that the BhYAB4b gene may regulate the development of leaves and fruits.

Key words: wax gourd, YABBY, gene cloning, expression analysis, subcellular localization, heterologous overexpression

WANG Baochen, LUO Chen, YAN Jinqiang, CHENG Xiaoxin, MO Renlian, LIU Wenrui, ZHANG Yuyang, JIANG Biao. Cloning，Expression，and Functional Research of BhYAB4b in Wax Gourd[J]. Acta Horticulturae Sinica, 2025, 52(9): 2353-2362.

Figures/Tables 7

Table 1 Primer sequences used for BhYAB4b gene in this study

引物用途Primer purpose	引物名称Primer name	引物序列（5′-3′）Primer sequence
基因克隆 Gene clone	Clone-BhYAB4b-F	ATGGTTTGGGGGGTGATCATTTACTTTTGT
基因克隆 Gene clone	Clone-BhYAB4b-R	TTATGTCAAAAATGAATGCCTTGGGAAACA
表达分析 Expression analysis	BhYAB4b-qRT-F	GCTCGAGCCTCCTTTCTGTT
表达分析 Expression analysis	BhYAB4b-qRT-R	GGTGTCACAGGAATCAAGCC
内参基因 Refenrence gene	BhUBQ-qRT-F	CCTAACTGGGAAGACGAT
	BhUBQ-qRT-R	CAAGACCAAGTGAAGGGT
	At-ACT-F	TAACAGGGAGAAGATGACTCAGATCA
	At-ACT-R	AAGATCAAGACGAAGGATAGCATGAG
亚细胞定位 Subcellular localization	BhYAB4b-pS1300-F	TACACCAAATCGACTCTAGTCTAGAAT GGTTTGGGGGGTGATCATTTACT
亚细胞定位 Subcellular localization	BhYAB4b-pS1300-R	CATGGTACCGGATCCACTAGTTGTCAA AAATGAATGCCTTGGGAAACA
拟南芥过表达 Overexpression in Arabidopsis	BhYAB4b-PCY-F	GAACACGGGGGACGAGCTCGGTACC ATGGTTTGGGGGGTGATCATTTACT
拟南芥过表达 Overexpression in Arabidopsis	BhYAB4b-PCY-R	GGTGGTGGTGGTGGTCGACGGATCCT TATGTCAAAAATGAATGCCTTGGGAAA

Fig. 1 Fragment amplification of BhYAB4b

Fig. 2 BhYAB4b protein hydrophilicity analysis（A）and prediction of secondary structure（B）

Fig. 3 Phylogenetic tree of wax gourd BhYAB4b protein and YAB4 proteins from other species The phylogenetic tree was constructed by the neighbor-joining method with 1 000 bootstrap replicates

Fig. 4 Subcellular localization of BhYAB4b protein

Fig. 5 Expression levels of BhYAB4b in different tissues（A）and fruits at different developmental stages（B）of B227 Different letters indicate significant differences（P < 0.05），the same below

Fig. 6 Phenotypic observation of 35S：BhYAB4b overexpression Arabidopsis A-C：Phenotypes of whole plants（A），rosette leaves（B），and fruits（C）of wild-type（WT）and 35S：BhYAB4b plants at 30 days after sowing；D：Expression levels of BhYAB4b；E-H：Comparison of rosette leaf and fruit length and width，and fruit length-to-width ratio. Values represent mean ± standard deviation（n = 3）

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Cloning，Expression，and Functional Research of BhYAB4b in Wax Gourd

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