甜瓜LBD基因家族的鉴定和果实发育进程中的表达分析

doi:10.13560/j.cnki.biotech.bull.1985.2022-1264

摘要/Abstract

摘要：

侧生器官边界域（lateral organ boundaries domain, LBD）基因广泛存在于高等植物中，可以参与调控植物生长发育及逆境响应。通过对甜瓜LBD转录因子的鉴定分析，旨在揭示其在果实发育进程中的调控作用。通过生物信息学分析明确LBD转录因子在基因组中的成员，分别命名为CmLBD1-CmLBD32；采用多序列比对分析甜瓜LBD 结构域的结构特征；运用邻接法、极大似然法进行系统进化树分析；以薄皮甜瓜发育各个阶段的果实（花后5-40 d，每间隔5 d取一次样）为试验材料提取RNA，并通过实时荧光定量技术检测LBD成员的基因表达水平。在甜瓜基因组上共鉴定出32个甜瓜LBD家族成员，分为7大类，分布于12条染色体上；多序列比对分析表明，32个LBD基因均具有典型的LOB结构域。在果实发育进程中，LBD家族基因在不同时期均有表达，其中CmLBD2和CmLBD28随着果实发育进程表达水平逐渐升高，其余家族成员在果实中表达水平相对较低。亚细胞定位表明，CmLBD2蛋白定位于细胞膜上。CmLBD2基因表达量与果实硬度间相关性分析表明，CmLBD2在果实成熟进程中基因表达量与硬度的变化显著负相关；利用外源1-MCP进行果实处理发现，1-MCP可以抑制CmLBD2基因表达，表明CmLBD2基因表达可能受到乙烯的调控。综上，甜瓜LBD基因在果实发育进程中可能发挥不同的作用，其中CmLBD2可能参与乙烯调控相关的果实后熟软化。

关键词: 甜瓜, LBD, 转录调控, 1-MCP, 基因表达, 果实发育, 生物信息学

Abstract:

The lateral organ boundaries domain（LBD）genes are widely found in higher plants, and can participate in the regulation of plant growth, development and stress response. Identification and analysis of the LBD transcription factor in melon（Cucumis melo）is aimed to unravel its regulatory role in the process of fruit development. Bioinformatics was used to identify the members of LBD transcription factor in melon genome and named as CmLBD1-CmLBD32. Multiple-sequence alignment was used to analyze the structure characteristics of LBD-domain in the melon. Maximum-likelihood and neighbor-joining were to have phylogenetic tree analyzed. The expressions of LBD gene family during fruit development were deteted by RT-qPCR using extracted RNAs from the fruits（five developing and mature fruits were harvested at 20, 25, 30, 35 and 40 d after pollination）. Total 32 LBD genes were identified from melon genome, and they can be divided into 7 categories and were distributed on 12 chromosomes. The multiple-sequence alignment analysis demonstrated that there were typical LOB domains in 32 LBD genes. LBD family genes expressed in different stages of fruits, the expressions of CmLBD2 and CmLBD28 gradually increased with the processes of fruit development, while relatively low for rest family members. Subcellular localization suggested that CmLBD2 protein was located in the cell membrane. The correlation analysis indicated that there was a negative correlation between the CmLBD2 expression and the fruit firmness during fruit ripening. CmLBD2 expression was inhibited by 1-Methylcyclopropene（1-MCP）after the fruit was processed with 1-MCP, indicating that CmLBD2 gene expression may be regulated by ethylene. In sum, LBD genes in the melon may play different roles in fruit development process, and CmLBD2 may be involved in ethylene regulation related to fruit ripening and softening after harvest.

Key words: Cucumis melo, LBD, transcriptional regulation, 1-MCP, gene expression, fruit development, bioinformatics

陈强, 邹明康, 宋家敏, 张冲, 吴隆坤. 甜瓜LBD基因家族的鉴定和果实发育进程中的表达分析[J]. 生物技术通报, 2023, 39(3): 176-183.

CHEN Qiang, ZHOU Ming-kang, SONG Jia-min, ZHANG Chong, WU Long-kun. Identification and Analysis of LBD Gene Family and Expression Analysis of Fruit Development in Cucumis melo[J]. Biotechnology Bulletin, 2023, 39(3): 176-183.

图/表 6

图1 甜瓜LBD保守结构域比对

Fig. 1 Alignment of conserved domains of LBD family in Cucumis melo

图2 甜瓜与拟南芥LBD转录因子的系统进化分析

Fig. 2 Phylogenetic tree of LBD transcription factors in C. melo and Arabidopsis

图3 甜瓜LBD基因家族表达谱

Fig. 3 Expression profile of LBD gene family in C. melo

图4 CmLBD2亚细胞共定位（膜）

Fig. 4 Subcellular location of CmLBD2（membrane）

图5 CmLBD2在采后贮藏过程中的表达量与果实硬度的相关性 3次生物学重复和3次技术重复，标准误显示在柱形图上，小写字母表示不同样品之间的差异（P<0.05）。下同

Fig. 5 Correlation between the expressions of CmLBD2 and fruit firmness during postharvest storage There are 3 biological and technical replicates, the standard error is displayed on the bar graph, and lowercase letters indicate the difference among different samples. The same below

图6 1-MCP对薄皮甜瓜果实CmLBD2基因表达量的影响

Fig. 6 Effects of 1-MCP on the expression of CmLBD2 in fruit of C. melo

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