Application of arabidopsis glycosyl transferase UGT74D1 in catalytic synthesis of auxin sugar ester

A technology of auxin sugar ester and glycosyltransferase, applied in the application of glycosyltransferase, the application field of Arabidopsis thaliana glycosyltransferase UGT74D1 in catalyzing the synthesis of auxin sugar ester, which can solve undiscovered problems, Achieve the effect of low efficiency and huge economic benefits

Inactive Publication Date: 2013-10-16
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, after searching, the role of other Arabidopsis glycosyltransferases in catalyzing the synthesis of auxin sugar esters has not been found, especially the report on the application of Arabidopsis thaliana glycosyltransferase UGT74D1 in the catalytic synthesis of auxin sugar esters

Method used

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  • Application of arabidopsis glycosyl transferase UGT74D1 in catalytic synthesis of auxin sugar ester
  • Application of arabidopsis glycosyl transferase UGT74D1 in catalytic synthesis of auxin sugar ester
  • Application of arabidopsis glycosyl transferase UGT74D1 in catalytic synthesis of auxin sugar ester

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Embodiment 1

[0028] Example 1 Cloning, prokaryotic expression and enzyme protein purification of Arabidopsis thaliana glycosyltransferase gene UGT74D1

[0029] 1. Cloning of Arabidopsis thaliana glycosyltransferase gene UGT74D1

[0030] The glycosyltransferase gene UGT74D1 involved in the present invention is cloned from Arabidopsis thaliana by RT-PCR amplification technology. Firstly, RNA was extracted from the young leaves of Arabidopsis thaliana by TRIzol method, and then the coding region cDNA of the gene was amplified by RT-PCR method. A pair of primers used for amplification are: UGT74D1-a: 5'-cgccatatgggagagaaagcgaaagc-3'; UGT74D1-b: 5'-ccgctcgagttacctcacaattttagc-3'. The RT-PCR amplification program is: 94°C (pre-denaturation), 5min; 94°C (denaturation), 10s; 55°C (annealing), 15s; 72°C (extension), 2min; 35cycle; 72°C (final extension), 10min. The amplified product is recovered and purified. The amplified target gene UGT74D1 was ligated with the intermediate vector pBluescript...

Embodiment 2

[0039] Example 2 Arabidopsis thaliana glycosyltransferase UGT74D1 catalyzes the synthesis of auxin sugar esters

[0040] 1. Enzyme-catalyzed reaction of auxin

[0041] The fusion protein of the purified Arabidopsis thaliana glycosyltransferase UGT74D1 (GST-UGT74D1, which can reflect the catalytic activity of UGT74D1, which is a common practice) was used for in vitro enzyme-catalyzed reactions, and six auxin compounds were selected as substrates, including Indole carboxylic acid (ICA), indole acetic acid (IAA), indole propionic acid (IPA), indole butyric acid (IBA), naphthalene acetic acid (NAA), 2,4-dichlorophenoxyacetic acid (2,4- D). UDP-glucose was used as the sugar donor in the catalytic reaction. The enzymatic reaction system is as follows:

[0042]

[0043] Put the above mixed reaction system in a constant temperature water bath at 37°C and react for 3 hours. The reaction was then terminated by adding 20 μl of trichloroacetic acid at a concentration of 240 mg / ml. ...

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Abstract

The invention discloses application of arabidopsis glycosyl transferase UGT74D1 in catalytic synthesis of auxin sugar ester. The amino acid sequence of the glycosyl transferase UGT74D1 is shown as SEQIDNo.2, and auxin comprises indolecarboxylic acid, heteroauxin, indole propionic acid, indolebutyric acid, naphthylacetic acid, and 2, 4-dichlorophenoxya-cetic acid. Putting the glycosyl transferase UGT74D1 and any one in the auxin into a reaction tube to be subjected to enzymatic reaction, and the corresponding auxin sugar ester can be catalyzed and synthetized. The application of the arabidopsis glycosyl transferase UGT74D1 in the catalytic synthesis of the auxin sugar ester provides a feasible method for synthetizing the sugar ester contents of the auxin matters by utilizing an enzymatic method in an efficient and exclusive mode and can bring huge economic benefits to auxin sugar ester synthesis industries.

Description

technical field [0001] The invention relates to the application of a glycosyltransferase, in particular to the application of an Arabidopsis thaliana glycosyltransferase UGT74D1 in catalyzing the synthesis of auxin sugar ester; it belongs to the field of biotechnology. Background technique [0002] Auxin is one of the important hormones of plants, which plays an important role in regulating and controlling the growth and development of plants. Naturally occurring auxins such as indole acetic acid (IAA), indole butyric acid (IBA), and synthetic auxins such as 2,4-dichlorophenoxyacetic acid (2,4-D), naphthalene acetic acid (NAA ) are widely used in scientific research and agricultural production. These auxins all contain a carboxyl group (-COOH). In plants, the hydroxyl group on the carboxyl group can be covalently bonded with glucose to form sugar esters of auxin. Sugar esters of auxin are considered to be inactive and storage forms of auxin (Bartel, 1997). At present, su...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P19/18
Inventor 侯丙凯金尚卉
Owner SHANDONG UNIV
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