豌豆VQ全基因组鉴定及其高温响应功能研究

doi:10.16420/j.issn.0513-353x.2025-0629

摘要/Abstract

摘要：

VQ基因家族在植物生长发育和逆境响应中具有重要功能。通过生物信息学方法鉴定豌豆中的VQ基因家族成员，分析其理化特性、进化关系、基因结构、启动子元件和共线性。结合RNA-seq与qRT-PCR解析表达模式，通过拟南芥遗传转化验证基因功能。本研究中鉴定出28个豌豆VQ基因，亚细胞定位主要在细胞核中，系统进化分为3个亚家族；基因结构以单外显子为主，启动子富含光响应、激素响应、胁迫响应和生长发育响应等多种元件；豌豆VQ基因存在4对物种内共线性基因对，与其他5个物种分别有18、15、19、36、76个共线基因对，进化上较为保守。该家族成员表达模式具有组织特异性，且部分成员响应高温胁迫。功能验证表明，过表达PsVQ15的拟南芥根长显著增长；过表达PsVQ15和PsVQ26均显著提升拟南芥在高温胁迫下的存活率与总叶绿素含量，其中过表达PsVQ26的拟南芥存活率最高、表型较好，表现出更强的耐热性。综上，豌豆VQ基因家族在植物生长发育和高温响应中发挥关键作用。

关键词: 豌豆, VQ基因家族, 全基因组鉴定, 生物信息学分析, 表达模式, 功能验证, 高温响应

Abstract:

The VQ gene family plays significant roles in plant growth，development，and stress response. In this study，members of the VQ gene family in pea（Pisum sativum）were identified through bioinformatics approaches，and their physicochemical properties，evolutionary relationships，gene structures，promoter elements，and synteny were analyzed. The expression patterns were analyzed by combining RNA-seq and qRT-PCR，and gene functions were verified through Arabidopsis genetic transformation. A total of 28 pea VQ genes were identified in this study，with subcellular localization mainly in the nucleus，and phylogenetically classified into three subfamilies. The VQ gene structure was predominantly single-exon，and the promoters were enriched with various cis-acting elements involved in light response，hormone response，stress response，and growth and development regulation. Four intraspecific syntenic gene pairs were identified in the pea VQ gene family，with 18，15，19，36，and 76 syntenic gene pairs detected between pea and the other five species，respectively，indicating a relatively conserved evolutionary pattern. Expression analysis revealed that pea VQ genes exhibited tissue-specific expression patterns，and certain members responded to high temperature stress. Functional verification demonstrated that，overexpression of PsVQ15 in Arabidopsis resulted in significantly longer roots，while both PsVQ15- and PsVQ26-overexpressing lines significantly elevated survival rates and total chlorophyll content under heat stress；the PsVQ26-overexpressing line exhibited the highest survival，superior phenotype，and thus stronger thermotolerance. In conclusion，the VQ gene family in pea plays a crucial role in plant growth，development，and heat response.

Key words: pea, VQ gene family, genome-wide identification, bioinformatics analysis, expression pattern, functional verification, heat stress response

中图分类号:

S643.3

李程勋, 徐晓俞, 李爱萍. 豌豆VQ全基因组鉴定及其高温响应功能研究[J]. 园艺学报, 2026, 53(9): 2639-2656.

LI Chengxun, XU Xiaoyu, LI Aiping. Genome-Wide Identification of the VQ Gene Family and Their Responses to Heat Stress in Pea[J]. Acta Horticulturae Sinica, 2026, 53(9): 2639-2656.

图/表 14

表1 本研究中所用引物序列

Table 1 Primers sequences used in this study

用途Purpose	引物名称Primer name	正向引物序列（5′-3′）Forward sequence	反向引物序列（5′-3′）Reverse sequence
基因的qPCR验证 Gene validation by qPCR	Actin-qPCR	CACAATTGGCGCTGAAAGAT	TAATTGAGTTAAATGTCGTCTCATGGAT
	PsVQ1-qPCR	GTGTCATCCACACTACGCCT	TCTCCCACCATCTCACTCGT
	PsVQ15-qPCR	GCTTTCCCGGGTTTGGTAGA	CAAAGGGGAAAACCCGGAGA
	PsVQ26-qPCR	TCCTGCGGAACCTTAACACC	TGATGAGAGGTTGTCGGTGC
	PsVQ28-qPCR	TCGTCTAACCCAAGCCCAAC	AACATCGTGGGTGTTTCCGA
基因克隆 Gene cloning	PsVQ15	GGGGTACCATGGATAAAGGCTCTGCA	TTCCGCGGCCGCTATGGCCGACGTCGACTCAAATTTCTCTCCACCG
基因克隆 Gene cloning	PsVQ26	GGGGTACCATGAACCTACCGGAATTT	TTCCGCGGCCGCTATGGCCGACGTCGACTTAGTCGAATATACTAAACAGATC
过表达载体验证 Validation of overexpression constructs	PsVQ15-1305a.1	GGGGGACATGGGTACCATGGATAA	TGCCTGCAGGTCGACTCAAAT
过表达载体验证 Validation of overexpression constructs	PsVQ26-1305a.1	CGGGGGACATGGGTACCATG	GCCTGCAGGTCGACTTAGTCGA

图1 豌豆VQ家族基因染色体定位和系统进化分析 A：豌豆VQ家族基因在染色体上的分布；B：豌豆（Ps）与拟南芥（At）VQ家族基因的系统进化树

Fig. 1 Chromosomal localization and phylogenetic analysis of the VQ gene family in pea A：Distribution of the VQ gene family on pea chromosomes；B：Phylogenetic tree of VQ genes from pea（Ps）and Arabidopsis（At）

图2 豌豆VQ家族基因结构和VQ蛋白保守基序示意图

Fig. 2 Gene structures of the VQ family and conserved motif composition of VQ proteins in pea

表2 豌豆VQ蛋白的保守基序信息

Table 2 Conserved motif information of VQ proteins in pea

基序 Motif	序列 Sequence	氨基酸数目 Number of amino acids	Pfam注释 Pfam annotation
Motif1	PTVVHTDASNFRAVVQKLTGS	21	VQ
Motif2	EEEEKAIKEKGFYLHPSPR	19	-
Motif3	MGCDENDFMVVAVPPLVDSDEQVVEPGGECSYEGFDEDVLLIPQMVENIW	50	-
Motif4	KPHPFRYDKQKHNINGPPPTPLMIRKPNSSHKQQRQPIIIY	41	-
Motif5	QHEGDETTSTTSSVEKEGVNSNINDDEQSPNNMMKFAEMPLFTPTSCDFF	50	-
Motif6	TNVTKSIKKRSRASKRAP	18	-
Motif7	GFKLYERRNSLK	12	-
Motif8	EILSPSILDFPSLVLSPVTPLIPDPFDR	28	-
Motif9	NLSGFDEAACENGDDGRNSFFISDLLLKECDLLHRELQPNNHFFF	45	-
Motif10	HQHHHQQQQQH	11	-

图3 豌豆VQ家族基因启动子顺式作用元件分析

Fig. 3 cis-Acting element analysis of the promoters of the pea VQ gene family

图4 豌豆VQ家族基因的物种内共线性关系

Fig. 4 Intraspecific syntenic relationships of the VQ gene family in pea

表3 豌豆VQ物种内共线性基因对的Ka/Ks值和分化时间

Table 3 Ka/Ks values and divergence time of intraspecific syntenic PsVQ gene pairs in pea

重复类型 Duplication type	重复基因1 Duplicated gene 1	重复基因2 Duplicated gene 2	Ka	Ks	Ka/Ks	分化时间/MYA Divergence time
片段重复 Segmental	PsVQ7	PsVQ21	0.30	2.87	0.10	95.52
	PsVQ1	PsVQ23	0.67	-	-	-
	PsVQ7	PsVQ27	0.25	0.90	0.28	29.92
	PsVQ16	PsVQ26	0.54	1.52	0.36	50.75

图5 豌豆与其他5个物种间VQ基因的共线性分析

Fig. 5 Syntenic analysis of VQ genes between pea and five other plant species

图6 豌豆与其他5个物种间VQ基因的进化分析 A：共线性VQ基因对Ks值的分布；B：共线性VQ基因对的Ks值；C：共线性VQ基因对的Ka值；D：共线性VQ基因对的Ka/Ks值。 Ps：豌豆；At：拟南芥；Ca：鹰嘴豆；Cc：木豆：Pv：菜豆；Gm：大豆

Fig. 6 Evolutionary analysis of VQ genes between pea and five other plant species A：Distribution of Ks values across the syntenic VQ gene pairs；B：Ks values of the syntenic VQ gene pairs；C：Ka values of the syntenic VQ gene pairs；D：Ka/Ks values of the syntenic VQ gene pairs Ps：Pisum sativum；At：Arabidopsis thaliana；Ca：Cicer arietinum；Cc：Cajanus cajan：Pv：Phaseolus vulgaris；Gm：Glycine max

图7 豌豆VQ家族基因在不同组织/器官中的表达热图

Fig. 7 Expression heatmap of the VQ gene family in different tissues/organs of pea

图8 豌豆VQ家族基因在高温胁迫下的表达热图 CK1：PR11-90对照；HS1：PR11-90高温胁迫处理；CK2：PR11-2对照；HS2：PR11-2高温胁迫处理

Fig. 8 Expression heatmap of the VQ gene family in pea under heat stress CK1：PR11-90 control；HS1：PR11-90 under heat stress；CK2：PR11-2 control；HS2：PR11-2 under heat stress

图9 4个差异表达基因的qRT-PCR与RNA-seq验证结果不同小写字母表示在P < 0.05水平下的差异显著性（n = 3）。下同

Fig. 9 Validation of 4 differentially expressed genes by qRT-PCR and RNA-Seq Different lowercase letters indicate the significant difference at P < 0.05 level（n = 3）. The same below

图10 PsVQ15和PsVQ26过表达载体构建与拟南芥遗传转化的电泳检测 A：PsVQ15和PsVQ26基因的PCR扩增产物；B：重组质粒转化大肠杆菌DH5α后的菌液PCR鉴定；C：重组质粒转化农杆菌GV3101后的菌液PCR鉴定；D：拟南芥T2代转基因植株的PCR鉴定。M：DL2000 Marker；WT：野生型

Fig. 10 Electrophoresis detection of overexpression vector construction and Arabidopsis genetic transformation for PsVQ15 and PsVQ26 A：PCR products of PsVQ15 and PsVQ26 gene；B：Bacterial PCR identification of recombinant plasmids transformed into E.coli DH5α；C：Bacterial PCR identification of recombinant plasmids transformed into Agrobacterium tumefaciens GV3101；D：PCR identification of T2 generation transgenic Arabidopsis plants. M：DL2000 marker；WT：Wild type

图11 过表达PsVQ15和PsVQ26拟南芥在高温胁迫下的表型分析 A：不同处理下拟南芥幼苗的生长状况；B：PsVQ15和PsVQ26过表达拟南芥植株的基因表达水平；C：不同处理下拟南芥幼苗的存活率、总叶绿素含量和根长。WT：野生型

Fig. 11 Phenotypic analysis of Arabidopsis overexpressing PsVQ15 and PsVQ26 under heat stress A：Growth performance of Arabidopsis seedlings under different treatments；B：Expression levels of PsVQ15 and PsVQ26 in the overexpressing plants；C：Survival rate，total chlorophyll content and root lengths of Arabidopsis seedlings under different treatments. WT：Wild type

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