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Jujube glutathione peroxidase gene

A glutathione peroxide and enzyme gene technology, applied in the field of genetic engineering, can solve the problems of no GPXs activity, limited understanding of function and structural characteristics, and achieve the effect of improving plant stress resistance and promoting understanding

Inactive Publication Date: 2012-07-04
AGRI BIOTECH RES CENT OF SHANXI PROVINCE +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, when some people studied citrus GPXs, they replaced cysteine ​​residues with selenocysteine ​​residues, but there was no activity similar to mammalian GPXs.
[0004] The structure, substrate specificity, and distribution of GPXs in plants are different. They play an important role in resisting external stress, but our understanding of their functions and structural properties is still very limited. need to do a lot of work

Method used

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  • Jujube glutathione peroxidase gene
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  • Jujube glutathione peroxidase gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Embodiment 1: jujube glutathione peroxidase gene wxya bioinformatics analysis of

[0037] The present invention constructs pot jujube ( Ziziphus jujuba Jujube glutathione peroxidase gene was screened from the cDNA library of fruiting branches of Mill hupingzao) wxya the full sequence.

[0038] will be measured wxya Search the NCBI nucleotide database for gene sequence through Blastx, and perform sequence similarity analysis; use DNAStar to compare the amino acid sequence and make the evolution tree of this protein; use ProtParam to calculate the relative molecular weight and theoretical isoelectric point of the protein; connect to http: / / www.expasy.org / prosite for conservative sequence analysis; submit the amino acid sequence of this protein to Swiss-Model to predict its tertiary structure.

[0039] Bioinformatics analysis showed that the cDNA sequence of jujube glutathione peroxidase gene was a full-length 510bp open reading frame (ORF). The sequence similar...

Embodiment 2

[0040] Example 2: wxya Construction of gene prokaryotic expression vector

[0041] 1. Materials, reagents and instruments

[0042] 1.1 Vectors and strains

[0043] Escherichia coli DH5α strain; the cloning vector pSPORT1-ZjGPX and the prokaryotic expression vector pGEX-4T-2 were all preserved by the Horticultural Crops Laboratory of Biotechnology Research Center, Shanxi Academy of Agricultural Sciences.

[0044] 1.2 Enzymes and kits

[0045] DNA recovery kit QIAquick? was purchased from QIAGEN; restriction endonuclease Bam H I. Sma I. Sal Ⅰ. T4 DNA ligase and DNA Marker (15000 bp, 2000 bp ladder) were purchased from Dalian TaKaRa (Bao Biological) Engineering Co., Ltd.

[0046] 1.3 Reagents and equipment

[0047] The antibiotic ampicillin was purchased from Shanghai Sangon Bioengineering Service Co., Ltd. Other reagents used in plasmid extraction and agarose gel electrophoresis were purchased from Tianjin Tianda Chemical Industry.

[0048] The main instruments an...

Embodiment 3

[0089] Example 3: wxya Expression and protein purification in Escherichia coli

[0090] 1 Materials, reagents and instruments

[0091] 1.1 Vectors and strains

[0092] Escherichia coli (Escherichia coli) BL21 (DE3) strain and prokaryotic expression vector pGEX-4T-2 were preserved by the Horticultural Crops Laboratory of Biotechnology Research Center, Shanxi Academy of Agricultural Sciences.

[0093] 1.2 Kits and instruments

[0094] Fusion protein purification kit Glutathione Resins was purchased from Dalian TaKaRa (Bao Biological) Engineering Co., Ltd.

[0095] The instruments used in this example mainly include ultrasonic instrument (West Germany Elma D-7700), DYY-6B electrophoresis instrument (Beijing Liuyi Instrument Factory), DYCZ-28A electrophoresis tank (Beijing Liuyi Instrument Factory), Sony digital camera (Sony (China) Co., Ltd. DSC-T500). For other instruments, see the Refined Molecular Biology Laboratory Guide.

[0096] 1.3 Reagent formulation

[0097] Re...

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Abstract

The invention relates to the technical field of genetic engineering, in particular to a jujube glutathione peroxidase gene. The nucleotide sequence and amino acid sequence of the gene are shown by SEQ ID NO:1 and SEQ ID NO:2 in the sequence table. In the invention, a plant expression vector is constructed, a transgenic arabidopsis plant is obtained, and the salt tolerance and drought tolerance ofthe plant are evaluated to prove that the stress resistance of the plant is improved by transferring the gene into the plant as a target gene, thereby being of great practical significance to the plant variety improvement; and further study on the stress resistance mechanism of the plant with good resistance is conducive to making clear the molecular structure, expression, function and control ofthe adversity related gene, provides certain experiment data and lays theoretical foundation for further sufficiently using the good-resistance genes to improve the stress resistance of the plant, ishelpful to understanding the effect of GPXs in the signal transduction of plant active oxygen, and promotes understanding of people about the active oxygen serving as a signal molecular.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to a jujube glutathione peroxidase gene. Background technique [0002] Mills first discovered that GPX, which has antioxidant effects in animal cells, is believed to be a defense against hemolysis caused by red blood cell oxidation, and named it glutathione peroxidase. In the past, it was considered to be the classic GPX, and now the enzyme with this function is named GPX-1. Later, the Flohe research group purified and analyzed the enzymes distributed in cells and acting on various organic peroxides and H 2 o 2 GPX, studies have shown that it is a tetramer composed of 4 protein subunits, each subunit contains 1 selenocysteine, and it forms the oxidation of the enzyme in the form of ionized selenol Restore the active center. A large number of studies have shown that mammalian GPXs have high catalytic activity and can quickly remove excess reactive oxygen species (ROS)...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/53C12N9/08C12N15/84A01H5/00
CPCC12Y111/01009C12N9/0065C12N15/8273A01H5/00
Inventor 曹秋芬孟玉平张晓娟张春芬曹尚银薛华柏
Owner AGRI BIOTECH RES CENT OF SHANXI PROVINCE
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