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A method for cultivating high nodulation nitrogen-fixing plants

A plant and nodulation technology, which is applied in the field of genetic engineering, can solve the problems of unseen regulatory effects, and achieve the effect of improving the ability of nodulation to fix nitrogen and increase soybean yield

Active Publication Date: 2020-08-18
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Haven't seen any related NRT1.2 Report on whether it plays a regulatory role in soybean root and root nodulation and symbiotic nitrogen fixation

Method used

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  • A method for cultivating high nodulation nitrogen-fixing plants
  • A method for cultivating high nodulation nitrogen-fixing plants
  • A method for cultivating high nodulation nitrogen-fixing plants

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Embodiment 1, GmNRT1.2 Analysis of expression patterns of genes in response to nitrate treatment.

[0022] 1. Material acquisition: The material used in the experiment is Williams 82 (referred to as W82); the material is carried out according to the following process: soybean seeds are sterilized with 70% sprinkler for 30 sec, sown in the vermiculite substrate soaked in low-nitrogen nutrient solution, They were cultured in a culture room with 16 h light / 8 h dark, light intensity 7000 LUX, temperature 26 °C, and relative humidity 70%. 15 days after sowing, the vermiculite was poured out, the soybean seedlings were soaked in a high-concentration nitric acid nutrient solution, and the main roots were harvested after 1 min, 5 min, 15 min, 1 h, 3 h, 24 h and 72 h of treatment, respectively. and lateral root samples. Among them, low nitrogen: 0.25 mM; high nitrogen: 15.75 mM.

[0023] 2. Isolation of mRNA: Trizol method was used to extract soybean total RNA. ①Put the tiss...

Embodiment 2

[0029] Embodiment 2, GmNRT1.2 Analysis of expression patterns of genes in response to rhizobia infection.

[0030] 1. Material acquisition: The material used in the experiment is Williams 82 (referred to as W82); the material is carried out according to the following process: soybean seeds are sterilized with 70% sprinkler for 30 sec, sown in the vermiculite substrate soaked in low-nitrogen nutrient solution, They were cultured in a culture room with 16 h light / 8 h dark, light intensity 7000 LUX, temperature 26 °C, and relative humidity 70%. 15 days after sowing, do inoculation with rhizobia, and each plant is inoculated with bradyrhizobia USDA110 bacterial solution (OD 600=0.08) 30 mL, and soybean roots were collected at 0, 1, 3, 5, 10 and 28 days after inoculation.

[0031] 2. Isolation of mRNA: Trizol method was used to extract soybean total RNA. ①Put the tissue into the grinder and grind it with liquid nitrogen for 3 times, then add 0.1-0.2 g of the ground tissue into a...

Embodiment 3

[0037] Example 3. Cultivation of transgenic soybeans with high nodulation nitrogen fixation ability.

[0038] 1. Extraction of total RNA from root tissue of Wilimas82 soybean.

[0039] The mortar has been treated at 180 ℃ for 8 hours or burned to eliminate RNase contamination; reagents such as chloroform, isopropanol, and ethanol should be freshly opened and uncontaminated; other equipment such as pipette tips, centrifuge tubes and reagents such as ultrapure water , NaAc, were treated with 1 ‰ DEPC water overnight, and then sterilized at 121 ℃ for 30 minutes, and the pipette tip and centrifuge tube were dried at 65 ℃ for later use; Trizol method was used to extract soybean total RNA:

[0040] (1) Take 100 mg of soybean root material and grind it with liquid nitrogen, add 1 mL of TRI pure reagent, after fully homogenizing, suck the homogenate into a 1.5 mL centrifuge tube, and place it at room temperature for 5 min;

[0041] (2) Add 200 μl chloroform, vortex to mix, let stand ...

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Abstract

The invention discloses a method for culturing a plant with high nodulation and nitrogen fixation capacity. A nucleotide sequence shown as SEQ ID NO: 1 in a sequence table is overexpressed in a plant,so as to obtain a transgenic plant with higher nodulation and nitrogen fixation capacity than a normal plant. An experiment proves that a receptor soybean plant is transformed by an overexpression vector constructed with the method, so as to obtain the transgenic plant, so that the nodulation and nitrogen fixation capacity of soybeans can be remarkably improved, and the significant practical meaning is realized for improvement of the yield of the soybeans.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to a method for cultivating transgenic plants. Background technique [0002] Soybean is an important grain and oil economic crop. Through root nodules, nitrogen in the air can be converted into ammonia nitrogen that can be absorbed and utilized by plants, thus providing the necessary nitrogen nutrition for soybean growth and development. The symbiotic nitrogen fixation process mediated by root nodules not only affects the normal growth and yield of soybean, but also helps to save energy and reduce environmental chemical pollution. At present, improving the efficiency of symbiotic nitrogen fixation has become one of the important ways to increase soybean yield and ensure the sustainable development of agriculture. [0003] Analyzing the mechanism of regulation of soybean root nodulation and symbiotic nitrogen fixation at the molecular level lays an important foundation f...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/82A01H5/00A01H6/54A01H6/00
CPCC12N15/8261
Inventor 王幼宁朱林李霞
Owner HUAZHONG AGRI UNIV
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