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Glyphosate-resistant Fusion Gene, Encoding Protein and Application

A fusion gene and anti-glyphosate technology, applied in the fields of application, fusion polypeptide, genetic engineering, etc., can solve extensive problems

Inactive Publication Date: 2017-01-25
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Human beings have been controlling weeds since the beginning of agricultural production, but weed control at that time was only an extremely extensive primary labor

Method used

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  • Glyphosate-resistant Fusion Gene, Encoding Protein and Application
  • Glyphosate-resistant Fusion Gene, Encoding Protein and Application
  • Glyphosate-resistant Fusion Gene, Encoding Protein and Application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Embodiment 1 Construction of anti-glyphosate fusion protein expression vector

[0026] The fusion gene formed by fusion of EPSPS gene CP4 and glyphosate N-acetyltransferase gene GAT is named CP4-GAT gene, and the encoded protein polypeptide sequence is shown in SEQ ID NO:3. The fusion gene formed by fusion of EPSPS gene CP4 and glyphosate oxidase gene GOX is named as CP4-GOX gene, and the encoded protein polypeptide sequence is shown in SEQ ID NO:4.

[0027] CP4-GAT gene (nucleotide sequence as shown in the 224-2053 bases in SEQ ID NO: 1) and the chloroplast transit peptide CTP at the 5' end of the gene (nucleotide sequence as in: SEQ ID NO: 1 Bases 11-223) and a terminator sequence at the 3' end (bases 2054-2259 in SEQ ID NO: 1), the gene is artificially synthesized, and BamHI restriction is set at the 5' end site, a KpnI restriction site is set at the 3' end, and the sequence is shown in SEQ ID NO:1. CP4-GOX gene (nucleotide sequence as shown in the 224-2898th base ...

Embodiment 2

[0034] Embodiment 2, transformation of rice

[0035] The method of obtaining transgenic rice is to adopt the existing technology (Lu Xiongbin, Gong Zuxun (1998) Life Science 10: 125-131; Liu Fan et al. (2003) Molecular Plant Breeding 1: 108-115). The mature and plump seeds of "Xiushui-134" were dehulled, and callus was induced as transformation materials. The Agrobacterium plates of the vectors pCambia1300-pUBI-CP4-GAT-p35S-1174 and pCambia1300-pUBI-CP4-GOX-p35S-1174 constructed in Example 1 were respectively taken. Pick a single colony to inoculate and prepare Agrobacterium for transformation. Put the callus to be transformed into the Agrobacterium bacterium solution with an OD of about 0.6 (preparation of the Agrobacterium bacterium solution: inoculate the Agrobacterium into the medium, and cultivate it at 28°C until the OD is about 0.6; the composition of the medium: 3g / L K 2 HPO 4 , 1g / LNaH 2 PO 4 , 1g / LNH 4 Cl, 0.3g / L MgSO 4 ·7H 2 O, 0.15g / L KCl, 0.01g / L CaCl 2...

Embodiment 3

[0036] Embodiment 3, fusion protein transgenic rice glyphosate resistance analysis

[0037] The T0 generation plants of the transgenic rice plants prepared by the method in Example 2 were transplanted into the greenhouse, and the herbicide resistance of the transgenic rice plants and the non-transgenic recipient control plant "Xiu Shui-134" were compared and analyzed. We obtained 201 transgenic lines (named CGAT) transgenic with pCambia1300-pUBI-CP4-GAT-p35S-1174 vector and 230 transgenic lines transgenic For CGOX) carry out different concentration glyphosate resistance determination, resistance effect is as shown in table 1:

[0038] Table 1*:

[0039] 2x 4x 6x Number of CGAT-resistant transformation events 201 178 92 Number of CGOX resistance transformation events 230 211 118

[0040] *Note: Table 1 shows the resistance test of rice plants sprayed with different concentrations of glufosinate-ammonium 15 days after sowing. The lines with no...

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Abstract

The invention discloses a glyphosate-resistant fusion gene, coding protein and application thereof, the fusion gene consists of the EPSPS protein encoding gene and the glyphosate N-acetyltransferase or glyphosate oxidase encoding gene; the EPSPS protein is one of the following: CP4, aroA, G7 or G10; glyphosate N-acetyltransferase encoding gene is GAT, and glyphosate oxidase encoding gene is GOX. Compared with glyphosate-resistant protein, the glyphosate-resistant fusion gene and the coding protein have the following advantages: the glyphosate-resistant fusion gene and the coding proteincan be resistant to glyphosate by two different tolerance mechanisms; two glyphosate tolerance mechanisms and high glyphosate resistance of transgenic plants can be conferred by one gene. The glyphosate-resistant fusion protein can be used in the anti-glyphosate of monocotyledonous and dicotyledonous plants, and is mainly used for herbicide-resistant insect-resistant maize, rice, soybean, wheat and rape.

Description

[0001] (1) Technical field [0002] The invention relates to a glyphosate-resistant fusion gene with two different glyphosate tolerance mechanisms, a fusion protein encoded by the gene, and an application of the fusion protein. [0003] (2) Background technology [0004] Weeds are the most important biological factors affecting yield in agricultural production. How to reduce the cost of weed control, improve the control efficiency, and reduce the environmental pollution during the control process are the most critical issues in agricultural science research. [0005] Weeds are produced along with human production activities, and their existence is the result of long-term adaptation to climate, soil, crops, farming systems and social factors and competition with cultivated crops. Human beings have been controlling weeds since the beginning of agricultural production, but weed control at that time was only an extremely extensive primary labor. With the development of science, e...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/62C07K19/00C12N15/54C12N15/53C12N15/82A01H5/00
CPCC12N9/1092C07K2319/00C12N9/1025C12N15/8275C12Y205/01019
Inventor 张先文王东芳沈志成
Owner ZHEJIANG UNIV
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