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NAC membrane-bound transcription factor gene gmntl1 in soybean Williams 82 and its application

A technology of transcription factor and membrane binding, applied in application, genetic engineering, plant genetic improvement and other directions, can solve the problem of no soybean NAC membrane-bound transcription factor, etc., to improve the salt stress tolerance and salt tolerance of plants. Effect

Active Publication Date: 2018-08-28
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

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Method used

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  • NAC membrane-bound transcription factor gene gmntl1 in soybean Williams 82 and its application
  • NAC membrane-bound transcription factor gene gmntl1 in soybean Williams 82 and its application
  • NAC membrane-bound transcription factor gene gmntl1 in soybean Williams 82 and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Example 1. Cloning of GmNTL1

[0035] 1.1 Extraction of Williams 82 total RNA

[0036] (1) Put Williams 82 plant material in a mortar and grind it into powder with liquid nitrogen (apply directly to the following experiment or freeze it in an ultra-low temperature refrigerator at -80℃ for later use);

[0037] (2) After the liquid nitrogen evaporates, immediately transfer 100-200mg of plant powder to a 1.5ml centrifuge tube, then quickly add 1ml of Trizol extract, vortex and shake to make the sample fully dissolved in the extract, and leave it at room temperature for 5 minutes;

[0038] (3) Centrifuge at 4°C at 12,000 rpm for 10 minutes, transfer 0.9ml of supernatant to a new 1.5ml centrifuge tube, add 0.2ml of chloroform, shake vigorously and mix for 15 seconds, and leave at room temperature for 2-5 minutes;

[0039] (4) Centrifuge at 4°C, 12,000 rpm, for 10 min, transfer 0.4ml of supernatant to a new 1.5ml centrifuge tube, add 0.4ml of isopropanol, turn up and down 15 times to m...

Embodiment 2

[0142] Example 2. Functional verification of the expression of Williams 82NAC membrane-bound transcription factor gene GmNTL1 in Arabidopsis

[0143] 2.1 Transformation of Arabidopsis thaliana by flower infection

[0144] (1) When Arabidopsis (Col-0 wild type) grows to 1cm bolting, the top is reduced to induce the formation of lateral inflorescence;

[0145] (2) One day before the transformation, take 1ml of activated Agrobacterium GV3101 containing the expression vector plasmid and add it to 40ml YEP medium containing the corresponding antibiotics and 50μg / ml rifampicin, and shake culture to OD at 28°C. 600 About 1.0-1.2;

[0146] (3) Centrifuge at 4,200 rpm for 10 min at room temperature, collect the bacteria, and resuspend the bacteria with the dip solution (5% sucrose, 0.05% Silwet L-77) to make the OD 600 About 0.8;

[0147] (4) Use a pipette to drop the Agrobacterium onto the inflorescences for infestation. After all the inflorescences are infected, place the Arabidopsis thaliana ...

Embodiment 3

[0185] Example 3. Functional verification of the Williams 82NAC transcription factor gene GmNTL1 expressed in soybean

[0186] 3.1 Soybean germination embryo vacuum infiltration assisted foreign gene transformation method to transform soybean

[0187] (1) Seed disinfection and pre-culture treatment

[0188] Sterilize soybean seeds with 70% ethanol for 5 minutes, and use 0.1% HgCl after removing the ethanol 2 Sterilize for 10 minutes, rinse with sterile water 5-6 times, soak seeds in sterile water at 25°C-28°C for 12 hours.

[0189] Put the soybeans whose embryos have expanded and germinated after soaking into the pre-culture medium, and cultured for 1 d at 28°C. The formula of the pre-culture medium is MS+3.0mg / L 6-BA(6-benzylamino adenine)+ 20g / L sucrose+7g / L agar, pH 5.8.

[0190] (2) After the seed embryo expands and germinates, it is cut to expose or damage the part of the germinating embryo

[0191] When the pre-cultured soybean embryo expands as a whole and the radicle grows to 0....

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Abstract

The invention discloses a NAC membrane binding transcription factor gene GmNTL1 in soybean Williams 82 and a plant expression vector containing the gene GmNTL1. The invention also discloses the application of the gene GmNTL1 to improve salt stress tolerance in Arabidopsis and soybean plants. Experiments have proved that the salt tolerance of transgenic plants of the present invention has been greatly improved compared with non-transgenic plants, which provides a theoretical basis and a practical basis for improving crop salt tolerance, and indicates that the genes of the present invention can be widely used. for breeding salt-tolerant plant varieties.

Description

Technical field [0001] The invention belongs to the technical field of biological genetic engineering, and in particular relates to a NAC membrane-bound transcription factor gene-GmNTL1 in soybean (Glycine max (L.) Merr) Williams 82 (Williams 82) and its application. Background technique [0002] Plants are exposed to the natural environment and are often affected by various environmental stresses, such as drought, extreme temperature, nutrient deficiency, and pests and diseases. These environmental stresses not only restricted the growth and development of plants, but also led to the reduction of food crops. The growth and development of higher plants and their response to environmental changes are achieved by regulating the expression of target genes. The interaction between transcription factors and gene cis-acting elements can be used as molecular switches for gene expression regulation. It is the interaction of transcription factors and cis-acting elements in the promoter ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/82A01H5/00A01H6/20A01H6/54
CPCC07K14/415C12N15/8273
Inventor 向凤宁王楠
Owner SHANDONG UNIV
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