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Method for cultivating transgenic nitrogen-fixing plants

A transgenic and plant technology, applied in the field of genetics and genetic engineering, can solve the problem of unclear function of miR172c, and achieve the effect of enhancing nitrogen fixation ability

Inactive Publication Date: 2011-10-26
INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although there are relatively many studies on miR172 at present, they mainly focus on the research on the aboveground part, while the underground function, especially the function of miR172c in soybean, has not been reported in detail, and the function of miR172c in root nodule development and nitrogen fixation is not clear.

Method used

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  • Method for cultivating transgenic nitrogen-fixing plants
  • Method for cultivating transgenic nitrogen-fixing plants
  • Method for cultivating transgenic nitrogen-fixing plants

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1: Analysis of tissue expression characteristics of gma-miR172c in soybean plants

[0021] 1. Real-time fluorescent quantitative PCR of gma-miR172c: Analysis of the expression characteristics of gma-miR172c in different soybean tissues by real-time fluorescent quantitative PCR technology

[0022] (1) Material acquisition, the material used in the experiment is Wilimas82 (Williams 82, hereinafter referred to as W82), and the material is processed according to the following process:

[0023] Soybean seeds are sterilized with 70% alcohol for 30 seconds, ddH 2 O washed 6 times and sowed in culture dishes covered with sterile filter paper, ddH 2 Soaked in O, germinated in the dark at 28°C for 3 days;

[0024] When the sprouts grow to 2cm, move them to a test tube with wet filter paper to accelerate the germination, so that the bean sprouts grow straight. When the sprouts grow to 4-5cm, move them to the activated rhizobium USDA110 bacterial solution (OD: 0.08) , in...

Embodiment 2

[0080] Example 2: A method for cultivating soybeans with high nitrogen fixation capacity based on the function of gma-miR172c

[0081]1. Extraction of total RNA from functional root nodule tissue of Wilimas82 soybean

[0082] The mortar has been treated at 180°C 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 all treated with 1‰ DEPC water overnight, then sterilized at 121°C for 30 minutes, and the pipette tip and centrifuge tube were dried at 65°C for later use; Trizol method was used to extract soybean total RNA:

[0083] (1) Take 50 mg of material (leaves) and grind the material with liquid nitrogen, add 1ml of redzol reagent, after fully homogenizing, inhale the homogenate into a 1.5ml centrifuge tube, and place it at room temperature for 5 minutes;

[0084] (2) A...

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Abstract

The invention discloses a method for cultivating transgenic nitrogen-fixing plants. The method comprises the following steps: constructing recombinant expression vectors by using nucleotide sequences shown in SEQ ID NO: 3 and vectors with tobacco cauliflower mosaic virus 35S promoters; then importing the recombinant expression vectors into target plants so as to obtain transgenic plants with enhanced nitrogen-fixing capacity as compared with normal plants. In the method disclosed by the invention, based on the new functions of gma-miR172c found by the applicant, the gma-miR172c is overexpressed in target plants by using the genetic engineering method, thereby obtaining the transgenic plants with enhanced nitrogen-fixing capacity as compared with normal plants.

Description

technical field [0001] The invention relates to the fields of genetics and genetic engineering, in particular to a method for cultivating transgenic nitrogen-fixing plants using gma-miR172c. Background technique [0002] Soybean is an annual herbaceous plant belonging to the legume family Soybean, and it is an important food and oil crop widely cultivated in the world; not only that, but there are many nodules growing on the roots of soybeans, which can fix free nitrogen in the air into ammonia nitrogen, which contributes to the growth of soybeans. Provides nourishment for growth. The nitrogen produced by rhizobia can only absorb half of the soybean in its lifetime, and the remaining half remains in the soil for the next crop to use. Undergraduate crops and other economic crops provide nitrogen nutrition, activate mineral elements, and reduce disease and insect damage. Therefore, the symbiotic nitrogen fixation mediated by root nodules not only affects the normal growth an...

Claims

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Application Information

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IPC IPC(8): C12N15/113C12N15/82A01H5/00
Inventor 李霞邹艳敏王幼宁陈亮石磊李东晓王延伟
Owner INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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