榅桲4CL基因的克隆与生物信息学分析

doi:10.6048/j.issn.1001-4330.2021.06.010

新疆农业科学 ›› 2021, Vol. 58 ›› Issue (6): 1055-1063.DOI: 10.6048/j.issn.1001-4330.2021.06.010

• 园艺特产·贮藏保鲜加工·植物保护 • 上一篇下一篇

榅桲4CL基因的克隆与生物信息学分析

杨波¹, 车玉红², 郭春苗¹, 木巴热克·阿尤普¹, 吴津蓉², 杜鹃²

1.新疆农业科学院园艺作物研究所,乌鲁木齐 830091;
2.新疆农业职业技术学院,新疆昌吉 831100

收稿日期:2020-09-03 出版日期:2021-06-20 发布日期:2021-06-15
通信作者: 车玉红(1977-),女,新疆昌吉人,教授,硕士,研究方向为果蔬贮藏保鲜加工及食品检测,(E-mail)cheyuhongxj@163.com
作者简介:杨波(1982-),男,新疆昌吉人,副研究员,硕士,研究方向为新疆特色果树丰产栽培与资源育种,(E-mail)yangboyys@163.com
基金资助:
新疆维吾尔自治区自然科学基金(2018D01A45)

Cloning and bioinformation analysis of 4CL gene in Cydonia oblonga Mill

YANG Bo¹, CHE Yuhong^2*, GUO Chunmiao¹, Mubareke·Ayoupu¹, WU Jingrong², DU Juan²

1. Institute of Horticulture,Xinjiang Academy of Agricultural Sciences; Urumqi 830091, China;
2. Xinjiang Agricultural Professional Technology College; Changji Xinjiang 831100,China

Received:2020-09-03 Online:2021-06-20 Published:2021-06-15
Correspondence author: CHE Yuhong ( 1977-),female,Fresh preservation,processing and food detection of fruits and vegetables,(E-mail)cheyuhongxj@163.com

摘要/Abstract

摘要： 【目的】克隆榅桲果肉(Cydonia oblonga Mill)中4-香豆酸辅酶A连接酶(4-coumarate CoA ligase,4CL)基因,研究其序列特征,为该基因在榅桲果实发育过程中的作用和功能奠定基础。【方法】以榅桲果肉为试材,基于GenBank报道的近缘物种4CL基因cDNA序列,应用Primer Premier 5.0软件设计PCR扩增引物。提取榅桲总RNA,经反转录后合成cDNA,应用RT-PCR方法成功扩增出4CL基因片段并克隆到pMD18-T载体。通过DNAstar软件进行同源序列比对,ClustalX结合MEGA4.1软件构建系统进化树,采用Protparam在线程序分析蛋白质的理化性质,用DNAstar的Protean程序预测二级结构。【结果】榅桲4CL基因其开放阅读框(ORF)序列为1 710 bp,编码570个氨基酸,蛋白分子质量为68.4 kD,与其他物种序列同源性最高达91.99%,进化关系上与水密桃较近;榅桲4CL蛋白为亲水性蛋白,二级结构中以无规卷曲(Random coil)为主,占比达到44.27%。【结论】获得了榅桲4CL基因全长编码区序列,探明了该序列的结构特征。

关键词: 榅桲; 4CL; 克隆; 序列分析

Abstract: 【Objective】 To clone and study the sequence characteristics of the 4-coumarate CoA ligase (4CL) gene from the pulp of Cydonia oblonga Mill will lay a foundation of further study on the role and function of this gene in the development of quince fruit.【Methods】 The pulp of Cydonia oblonga Mill was used as test material and based on the 4CL gene cDNA sequence of related species reported by GenBank, primers were designed by primer premier 5.0 software. The total RNA was extracted and the cDNA was synthesized by reverse transcription. The 4CL gene fragment was amplified by RT-PCR and cloned into pMD 18-T vector. DNAstar software was used for homologous sequence alignment. ClustalX combined with MEGA 4.1 software was used to construct phylogenetic tree. Protparam online program was used to analyze the physical and chemical properties of proteins, and the protein program of DNAstar was used to predict the secondary structure.【Results】 The open reading frame (ORF) sequence of 4CL gene in Cydonia oblonga Mill was 1,710 BP, which encoding protein of 570 amino acid residues, with a predicted molecular mass of 68.4 kD and the highest homology with other species was 91.99%. The evolutionary relationship between the two genes was close to that of water dense peach. 4CL protein in Cydonia oblonga Mill was a hydrophilic protein, and the secondary structure of 4CL protein was mainly random coil, which was accounting for 44.27%.【Conclusion】 The full-length coding region of 4CL gene in Cydonia oblonga Mill was successfully obtained, and the structural characteristics of the sequence were confirmed.

Key words: Cydonia oblonga Mill; 4CL; clone; sequence analysis

中图分类号:

S188
S662.9

杨波, 车玉红, 郭春苗, 木巴热克·阿尤普, 吴津蓉, 杜鹃. 榅桲4CL基因的克隆与生物信息学分析[J]. 新疆农业科学, 2021, 58(6): 1055-1063.

YANG Bo, CHE Yuhong, GUO Chunmiao, Mubareke·Ayoupu, WU Jingrong, DU Juan. Cloning and bioinformation analysis of 4CL gene in Cydonia oblonga Mill[J]. Xinjiang Agricultural Sciences, 2021, 58(6): 1055-1063.

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榅桲4CL基因的克隆与生物信息学分析

Cloning and bioinformation analysis of 4CL gene in Cydonia oblonga Mill

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