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Ubiquitin-1 promoter-based plant aphid resistance implementation method

A promoter and plant technology, applied in the field of bioengineering, can solve the problems of easy pollution of the environment by pesticides, and achieve the effect of facilitating promotion and low cost

Inactive Publication Date: 2018-03-23
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Pesticides play an important role in resisting pests, however, pests are gradually developing resistance to pesticides, in addition, pesticides are also easy to pollute the environment

Method used

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  • Ubiquitin-1 promoter-based plant aphid resistance implementation method
  • Ubiquitin-1 promoter-based plant aphid resistance implementation method
  • Ubiquitin-1 promoter-based plant aphid resistance implementation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Cloning of EβF Gene of Peppermint

[0019] 1. Extraction of total RNA from the peppermint genome: Take the peppermint leaf tissue, grind it in liquid nitrogen, add it to a 1.5mL Eppendorf (EP) centrifuge tube filled with lysate, shake it fully, and follow the instructions of TIANGEN Biochemical Technology Co., Ltd. Total RNA was extracted according to the instructions of the company's plant total RNA kit. The quality of total RNA was identified by agarose gel electrophoresis, and then the RNA content was determined on a spectrophotometer.

[0020] 2. Cloning of the peppermint EβF gene: using the extracted total RNA as a template, cDNA was synthesized under the action of PowerScript reverse transcriptase; gene-specific primers were designed according to the sequence of the EβF gene, and amplified from the total cDNA by PCR EβF gene, and sequenced.

[0021] Through the above steps, the full-length 1650bp of peppermint EβF gene is obtained, its nucleotide coding sequence...

Embodiment 2

[0028] Construction of Plant Overexpression Vector Containing EβF Gene

[0029] The partial sequence of the EβF gene was constructed on the plant overexpression vector (pDE1005) using recombinase. In order to facilitate the construction of the expression vector, the restriction site of SpeI was introduced into the forward primer, and the restriction site of SacI was introduced into the reverse primer. point, the primers are as shown in Table 3, and its promoter is the ubiquitin-1 promoter;

[0030] Table 3 PCR primers constructed by pDE1005-EβF body

[0031]

Embodiment 3

[0033] Obtaining Transgenic Wheat Plants by Transforming Immature Embryos of Wheat with Biolistic Method

[0034] 1. Immature embryo culture: select young ears about 14 days after flowering, collect grains of appropriate size (immature embryo size is 1.0-1.5mm), and disinfect the surface with 70% ethanol for 30 seconds in an ultra-clean workbench, and disinfect with 10% sodium hypochlorite for 10 seconds. -12min, rinsed with sterile water 3 times, gently peeled off the immature embryos, inoculated in SD 2 Callus was induced on culture medium.

[0035] 2. Gene gun transformation of wheat immature embryos: inoculate the immature embryo calli of about 8 days as the recipient of exogenous genes. The plasmid DNA containing the target gene is wrapped with gold powder bullets for transformation. Before the bombardment of the gene gun, the receptors were placed in the hyperosmotic medium (SD 2 +0.4mol L-1 sorbitol) for 4 hours. After the bombardment, the callus was placed on the h...

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Abstract

The invention relates to a ubiquitin-1 promoter-based plant aphid resistance implementation method. An E-beta-farnesene synthase (EbetaFS) gene is cloned from peppermint by means of genetic engineering, and a ubiquitin-1 promoter is constructed for driving an EbetaF vector. An gene is transferred into wheat by using a gene gun transformation method; E-beta-farnesene is synthesized by catalyzing FPP in the wheat; the release of the E-beta-farnesene in transgenic wheat is realized by synthesizing an aphid alarm element E-beta-farnesene in the wheat, so that the aim of avoiding aphids is achieved.

Description

technical field [0001] The invention relates to a technology in the field of bioengineering, in particular to a method for enhancing plant aphid resistance based on ubiquitin-1 promoter driving EβF gene. Background technique [0002] Aphids are one of the main pests in agricultural production. The temperature rises during the wheat growth period in the main wheat planting areas of my country, and "warm winter" occurs, which leads to rampant aphid damage. Economic losses caused by aphids and the viruses they spread in wheat production every year Very serious. Since the main varieties of wheat in my country are not very resistant to aphids at present, how to effectively control aphids has become one of the major problems in wheat production. Pesticides play an important role in resisting pests, but pests are gradually developing resistance to pesticides. In addition, pesticides are easy to pollute the environment. Genetic modification has made a major contribution to helping ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/60C12N15/82A01H5/00A01H6/46
CPCC12N9/88C12N15/8216C12N15/8286C12Y402/03047
Inventor 唐克轩马嘉伟付雪晴张婷婷赵静雅谢利辉黎凌孙小芬
Owner SHANGHAI JIAO TONG UNIV
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