Corn chloroplast transgene expression vector containing herbicide resistant gene and application thereof

An expression vector, chloroplast technology, applied in the application, the use of vectors to introduce foreign genetic material, and the cells modified by the introduction of foreign genetic material, etc., can solve the problem that no new varieties of glyphosate resistance appear, and rarely reach the level of promotion, etc. problem, to achieve excellent glyphosate resistance, improve glyphosate resistance, and achieve the effect of small molecular weight

Inactive Publication Date: 2012-07-04
THE INST OF BIOTECHNOLOGY OF THE CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although some EPSPS genes resistant to herbicides have been cloned from microorganisms, plants, etc., and corresponding transgenic plants have been obtained, but few of them can reach the level of promotion
[0006] At present, there are no new glyphosate-resistant varieties that have been widely promoted in China.

Method used

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  • Corn chloroplast transgene expression vector containing herbicide resistant gene and application thereof
  • Corn chloroplast transgene expression vector containing herbicide resistant gene and application thereof
  • Corn chloroplast transgene expression vector containing herbicide resistant gene and application thereof

Examples

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Effect test

Embodiment 1

[0036] Example 1. Acquisition of EPSP synthase gene, construction and transformation of expression vector, identification of transgenic maize.

[0037] 1. Extraction of maize chloroplast genome:

[0038] (1) Remove the veins from 5-10g of corn leaves, cut the leaves into pieces, and grind them with liquid nitrogen;

[0039] (2) Add 5 times the volume of buffer A (buffer A: 50mM Tris-HCl, 20mM EDTA, 0.5M sucrose, 0.25M Vc, pH3.5) pre-cooled at 4°C, homogenate and filter the filtrate with 8 layers of medical gauze until pre-cooled In a 50mL centrifuge tube, discard the residue;

[0040] (3) The filtrate was centrifuged at 1500g / min for 15min, the supernatant was discarded, and the precipitate was retained;

[0041] (4) Add 3mL bufferA to the precipitate, fully suspend, and transfer to a 10mL centrifuge tube;

[0042] (5) Centrifuge at 2000g / min for 15min, discard the supernatant, and collect the precipitate;

[0043] (6) Add 2mL bufferB and suspend fully (buffer B: 50mM Tris...

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Abstract

The invention provides a corn chloroplast transgene expression vector containing herbicide resistant gene and application thereof. The transgene expression vector comprises an expression box of 5-enol form acetone shellokimic acid-3-phosphate synthase (EPSP) gene, a homologous recombination fragment atpB in a corn chloroplast gene group and a homologous recombination fragment rbcL, wherein the expression box of the EPSP synthase gene is positioned between the homologous recombination fragment atpB and the homologous recombination fragment rbcL. The invention further provides a construction method of the chloroplast transgene expression vector. The invention also provides application of the chloroplast transgene expression vector in improving corn glyphosate resistance, which comprises leading the expression vector to convert callus or cells of corns, and screening to obtain resistant callus or cells; and inducing resistant callus or cells to differentiate, and regenerating to achieve corn resistance plants. The corn chloroplast transgene expression vector containing herbicide resistant gene can remarkably improve corn glyphosate resistance and has the advantages of being high in biological safety, capable of saving time, efficient, little in energy consumption, low in cost and the like.

Description

technical field [0001] The present invention relates to a chloroplast expression vector, in particular to a maize chloroplast expression vector containing EPSP synthase (5-enol-pyruvylshikimate-3-phosphate synthase) gene and a construction method thereof. The present invention also relates to the EPSP synthase The application of an enzyme gene maize chloroplast expression vector in improving maize glyphosate resistance belongs to the field of maize glyphosate resistance transgenic engineering. Background technique [0002] In 1988, Boynton et al. bombarded atpB mutant Chlamydomonas with chloroplast DNA carrying the atpB wild-type gene for the first time by using the particle gun method to restore the ability of photosynthesis, thus marking the beginning of chloroplast genetic engineering. In 1900, the rrn16 gene was expressed in tobacco chloroplasts for the first time by using the gene gun method, which opened the curtain of chloroplast transformation in higher plants. Gras...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/82C12N5/10C12N15/66A01H5/00
Inventor 李轶女张志芳王金辉王国增程奇倪丕冲沈桂芳
Owner THE INST OF BIOTECHNOLOGY OF THE CHINESE ACAD OF AGRI SCI
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