不同成熟果色辣椒中<i>PSY1</i>基因的差异表达分析

doi:10.7668/hbnxb.20194772

摘要/Abstract

摘要：

为了研究PSY1基因在辣椒不同成熟果色形成过程中的作用机理,以Y15016、Y15016-2、SP01、SP02、Z1为材料,克隆PSY1基因并进行时空表达分析,结合部分生物信息学方法,对PSY蛋白功能性质及辣椒不同果色材料中PSY1基因的表达情况进行研究分析。结果表明:5个品种辣椒中均能克隆到PSY1全长基因,且序列无差异;基因结构分析表明,辣椒PSY1基因包含6个外显子、5个内含子,全长为2 844 bp,其CDS全长为1 260 bp,编码419个氨基酸。序列比对和进化树分析表明,辣椒PSY蛋白与同科的番茄、烟草同源PSY蛋白在亲缘关系上最为接近。qRT-PCR分析结果显示:PSY1基因在试验选用的5个品种辣椒根、茎、叶、花组织中均显示表达,在根组织中表达量最低,叶组织中的表达量最高;在橙色突变体Y15016-2的表达量总体高于其在野生型Y15016中的表达量,而在黄色突变体SP02中,其表达量明显低于野生型SP01;在果实不同发育阶段,PSY1基因除Ⅲ期表达有所降低外,其表达量随果实发育总体呈现上升趋势,且在不同果色材料的成熟期(Ⅳ—Ⅴ期)达到最大值。PSY1基因启动子分析结果显示,供试材料中其序列未见差异。结果表明,在不同果色辣椒的果色形成过程中,PSY1基因的差异表达可能起到了较为重要的作用。

关键词: 辣椒, 八氢番茄红素合成酶, 类胡萝卜素代谢途径, 基因克隆, 基因表达

Abstract:

In order to study the mechanism of PSY1 gene in different ripe fruit colors of peppers, Y15016,Y15016-2,SP01,SP02 and Z1 were used as materials,and the functional properties of PSY protein and the expression of PSY1 gene in different fruit color materials of pepper were studied and analyzed by combining some bioinformatics methods.The results showed that the full-length PSY1 gene could be cloned in all five pepper varieties,and there was no difference in sequence.Gene structure analysis showed that the PSY1 gene contained six exons and five introns with a total length of 2 844 bp,and its CDS contained 1 260 bp and encoded 419 amino acids.Sequence alignment and phylogenetic tree analysis showed that the PSY protein of pepper was the closest to the homologous PSY protein of tomato and tobacco of the same family.The results of qRT-PCR analysis showed that the expression level of PSY1 gene in the five materials was lowest in the root tissue and highest in the leaf tissue.Besides,the PSY1 gene was expressed in all the tissues of these materials.The expression level of orange mutant Y15016-2 was higher than that of wild-type Y15016,while that of yellow mutant SP02 was significantly lower than that of wild-type SP01.At different stages of fruit development,the expression of PSY1 gene increased with the development of fruits,except for the decrease in stage Ⅲ,and reached the maximum value at the maturity stage(Ⅳ—Ⅴ)of different fruit color materials.The results of PSY1 gene promoter analysis showed that there was no difference in the sequences in the test materials.The results suggest that the differential expression of PSY1 gene may play an important role in the formation of peppers with different fruit colors.

Key words: Pepper, Phytoene synthase, Carotenoid metabolic pathway, Gene cloning, Gene expression

陶功臣, 马玉杰, 文军琴, 王亚艺, 李全辉. 不同成熟果色辣椒中PSY1基因的差异表达分析[J]. 华北农学报, 2024, 39(3): 15-24. doi: 10.7668/hbnxb.20194772.

TAO Gongchen, MA Yujie, WEN Junqin, WANG Yayi, LI Quanhui. Differential Expression Analysis of PSY1 Gene in Peppers with Different Ripe Fruit Colors[J]. Acta Agriculturae Boreali-Sinica, 2024, 39(3): 15-24. doi: 10.7668/hbnxb.20194772.

http://www.hbnxb.net/CN/Y2024/V39/I3/15

图/表 12

图1 试验材料

Fig.1 Experimental material A.Y15016;B.Y15016-2;C.SP01;D.SP02;E.Z1。

图2 辣椒PSY1基因克隆及基因结构图 A.辣椒PSY1基因克隆琼脂糖凝胶电泳结果;B.辣椒PSY1基因结构图。

Fig.2 Cloning and gene structure chart of PSY1 gene in Capsicum annuum L. A.Agarose gel electrophoresis of PSY1 gene cloning in Capsicum annuum L.;B.The gene structure prediction of PSY1 in Capsicum annuum L..

图3 辣椒PSY1基因CDS全长及氨基酸序列

Fig.3 CDS full length and prediction amino acid sequence of PSY1 gene in Capsicum annuum L.

图4 辣椒PSY蛋白信号肽和跨膜结构域预测 A.辣椒PSY蛋白信号肽预测;B.辣椒PSY蛋白跨膜结构域预测。

Fig.4 Pepper PSY protein signal peptide and transmembrane domain prediction A.Protein signal peptide prediction of pepper PSY protein;B.Transmembrane domain prediction of pepper PSY protein.

图5 辣椒PSY氨基酸序列的亲水性和疏水性分析

Fig.5 PSY amino acid sequence hydrophilicity and hydrophobicity analysis in Capsicum annuum L.

图6 辣椒PSY蛋白二级及三级结构预测 A.辣椒PSY蛋白二级结构预测;B.辣椒PSY蛋白三级结构预测。

Fig.6 Prediction of PSY protein secondary and tertiary structure in Capsicum annuum L. A.Prediction of PSY protein secondary structure in Capsicum annuum L.;B.Prediction of PSY protein tertiary structure in Capsicum annuum L..

图7 辣椒PSY氨基酸序列保守域预测

Fig.7 Prediction of PSY amino acid sequence conservative domain in pepper

图8 辣椒PSY蛋白与其他植物同源PSY蛋白氨基酸序列多重比对

Fig.8 PSY protein multiple comparison of amino acid sequence between Capsicum annuum L. and other plants

图9 辣椒与其他物种PSY氨基酸序列的系统进化树

Fig.9 PSY proteins amino acid sequence phylogenetic tree of Capsicum annuum L. and other plants

图10 5个品种辣椒成熟期不同组织PSY1基因的相对表达水平不同小写字母表示PSY1基因在辣椒不同组织间表达差异显著(P<0.05);不同大写字母表示PSY1基因在辣椒不同品种间表达差异显著(P<0.05)。

Fig.10 PSY1 genes relative expression levels in different tissues of full-blown pepper during five varieties Different lowercases represent significant differences in PSY1 gene expression between tissue development(P<0.05);different uppercases represent significant differences in PSY1 gene expression between different varieties(P<0.05).

图11 5个品种辣椒果实不同时期PSY1基因的相对表达水平不同小写字母表示PSY1基因在辣椒果实发育过程表达差异显著(P<0.05);不同大写字母表示PSY1基因在辣椒不同品种间表达差异显著(P<0.05)。

Fig.11 PSY1 genes relative expression levels in different periods of pepper fruit in five varieties Different lowercases represent significant differences in PSY1 gene expression between fruit development(P<0.05);different uppercases represent significant differences in PSY1 gene expression between different varieties(P<0.05).

图12 辣椒PSY1基因启动子分析 A.Y15016、Y15016-2、SP01、SP02、Z1和参考基因组CM334中PSY1基因启动子序列比对;B.PSY1基因启动子顺式作用元件预测。

Fig.12 PSY1 gene promoter analysis in pepper A.Alignment of PSY1 gene promoter sequences in Y15016,Y15016-2,SP01,SP02,Z1 and reference genome CM334;B.PSY1 gene promoter cis-acting element prediction.

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不同成熟果色辣椒中PSY1基因的差异表达分析

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