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J. Hefei Univ. Tech. (Nat. Sci.)

合肥工业大学学报(自然科学版)

JOURNAL OF HEFEI UNIVERSITY OF TECHNOLOGY (NATURAL SCIENCE)

Competent Authorities: Ministry of Education of the People's Republic of China

Sponsor by: Hefei University of Technology

ISSN 1000-1646, CN 34-1001/N

Cloning and expression analysis of GOX1 gene in Arabidopsis thaliana

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拟南芥 GOX1 基因的克隆和表达分析

Journal Information

J. Hefei Univ. Tech. (Nat. Sci.), May.2023, Vol.46 No.5: 698-703

DOI: 10.3969/j.issn.1003-5060.2023.05.020

Authors

TIAN Mimi, MU Xiujie, ZHANG Ruihan, HAN Yi

(School of Food and Biological Engineering, Hefei University of Technology, Hefei 230601, China)

Abstract and Keywords

Abstract: Glycolate oxidase 1(GOX1) is one of the key enzymes in the photorespiration process of Arabidopsis thaliana, which can oxidize glycolic acid to glyoxylic acid and hydrogen peroxide. In this paper, wild-type Arabidopsis thaliana Columbia(Col-0) was used as the research material. The 1 104 bp CDS sequence of GOX1 gene was obtained by reverse transcription-polymerase chain reaction (RT-PCR) technology, its gene function was analyzed preliminarily using bioinformatics technology, PET28a(+)-GOX1 prokaryotic expression vector with His tag was transformed into E. coli competent BL21, and SDS-PAGE electrophoresis was used to detect the protein product after induced expression. The GOX1 gene encodes 367 amino acids, the relative molecular mass is 40 341.48, and the theoretical isoelectric point is 9.16. It is a hydrophilic protein and is located in peroxisomes. SDS-PAGE detected the band of interest at 40 kDa, which was consistent with the theoretical prediction. Through the analysis of GOX1 gene expression, it has laid the foundation for further exploration of protein function and regulation mechanism.

Keywords: glycolate oxidase 1(GOX1); gene cloning; bioinformatics analysis; prokaryotic expression

作者信息

田蜜蜜，穆秀杰，张瑞含，韩毅

（合肥工业大学食品与生物工程学院，安徽合肥230601）

摘要和关键词

摘要: 乙醇酸氧化酶1（glycolate oxidase 1，GOX1）是拟南芥光呼吸过程中的关键酶之一，可将乙醇酸氧化为乙醛酸和过氧化氢。文章以野生型拟南芥（Arabidopsis thaliana）Columbia（Col-0）为研究材料，通过反转录（reverse transcription，RT）和聚合酶链式反应（polymerase chain reaction，PCR）技术获得GOX1基因1104 bp的CDS序列，运用生物信息学技术初步分析其基因功能，并构建带有His标签的PET28a（+）-GOX1原核表达载体，转化至大肠杆菌感受态BL21中，采用SDS-PAGE电泳检测诱导表达后的蛋白产物。GOX1基因编码367个氨基酸，相对分子质量为40341.48，理论等电点为9.16，属于亲水性蛋白，定位于过氧化物酶体。SDS-PAGE在40 kDa处检测到目的条带，与理论值预测一致。通过对GOX1基因的表达分析，为进一步探究蛋白功能和调控机制奠定了基础。

关键词: 乙醇酸氧化酶1（GOX1）；基因克隆；生物信息学分析；原核表达

Funding

国家自然科学基金资助项目（31300225）

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