Improvement of Astragaloside content by endogenous gene overexpression technology

An overexpression and endogenous gene technology, applied in genetic engineering, plant gene improvement, recombinant DNA technology, etc., can solve the problems of long growth cycle of transgenic plants, more genes and less

Inactive Publication Date: 2007-05-09
SHANGHAI UNIV OF T C M
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  • Abstract
  • Description
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Problems solved by technology

However, there are few reports on the introduction of multiple genes, and they are mainly related to the resistance of crops to diseases and insect pests. In these reports, the foreign genes are directly introduced into the crops, and the transgenic crops are then grown in the field, so the obtained transgenic plants still need a long time. livespan

Method used

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  • Improvement of Astragaloside content by endogenous gene overexpression technology
  • Improvement of Astragaloside content by endogenous gene overexpression technology
  • Improvement of Astragaloside content by endogenous gene overexpression technology

Examples

Experimental program
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Effect test

Embodiment 1

[0024] 1) Total RNA extraction (one-step method)

[0025] Preparation of reaction reagents: denaturing solution (26mM sodium citrate pH 4.0, 0.5% sodium lauryl sarcosinate, 0.125M 2-mercaptoethanol, 4M guanidine isothiocyanate); NaAc (2M, pH 4.7); phenol : chloroform: isoamyl alcohol (25:24:1); 70% ethanol. All reagents are prepared with RNase-free water treated with 0.1% DEPC water, and all glass containers are RNase-free.

[0026] Operating procedures: Take an appropriate amount of fresh plant tissue and place it in a ceramic mortar, add liquid nitrogen and quickly grind it to a fine powder, transfer it to a 1.5ml centrifuge tube containing 650 μl denaturing solution and mix well; add 65 μl 2M NaAc (pH 4.0), and invert repeatedly Mix well; then add 650 μl phenol: chloroform: isoamyl alcohol, mix well, and ice-bath for 15 min; 4°C, 12000 rpm, centrifuge for 20 min; transfer the supernatant to a new 1.5 ml centrifuge tube, add an equal volume of isopropanol, mix After homoge...

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Abstract

The invention discloses an improvement of Astragaloside content by endogenous gene overexpression technology, wherein the two metabolism reaction coupled key enzyme genes of the polysaccharide metabolic pathway, i.e., ugp gene (uridine diphosphate glucose phosphorylase gene) and gbss gene (adenosine diphosphate glucose glycoside transferase gene) are modified in vitro, so as to reinforce the enhanced expression regulation element and construct intermediate carrier, and electrical perforation method is employed to transfer the expression intermediate carrier pBI-UG into Agrobacterium rhizogenes LBA-9402, thus obtaining polysaccharide synthesize double gene Radix Astragali hairyroot.

Description

Technical field: [0001] The invention relates to the technical field of traditional Chinese medicine. It specifically relates to a method for increasing the content of astragaloside IV by an endogenous gene overexpression technique. Background technique: [0002] Astragalus membranacius or A.mongholicus is one of the important Chinese herbal medicines in my country. It has the functions of tonifying and solidifying the exterior, diuresis and detoxification, promoting muscle and reducing sores, diuretic and swelling, and is the main component of many traditional Chinese medicine compounds, patent medicines and health care products. With the strengthening of people's growing health care concept, the demand for Astragalus membranaceus will increase year by year; but at present, the wild resources of Astragalus membranaceus are decreasing year by year, the quality of cultivated varieties is declining, and they are facing problems such as pesticide pollution and heavy metal resid...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/09C12N15/10C12N15/54C12N15/63C12N15/87C12N15/82C12N5/04C12N15/70C12N15/74A01H4/00A61K36/481A61P9/10
Inventor 杜旻胡之璧刘涤周吉燕王子艳倪跃元
Owner SHANGHAI UNIV OF T C M
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