Application of miR399 in promotion of absorption and utilization of nitrogen and phosphorus and blooming of soybeans

A mir399, 1.mir399 technology, applied in the field of genetic engineering, can solve the problems of unclear growth and development of leguminous plants, unclear relationship, limited practical application, etc., to promote the absorption and utilization of nitrogen and phosphorus, increase biomass, increase Effects of nitrogen and phosphorus content in soybean

Inactive Publication Date: 2017-01-11
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The direct relationship between miR399 and soybean nitrogen and phosphorus nutrient balance is still unclear, and the effect of miR399 on the growth and development of legumes is unclear, which limits its practical application

Method used

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  • Application of miR399 in promotion of absorption and utilization of nitrogen and phosphorus and blooming of soybeans
  • Application of miR399 in promotion of absorption and utilization of nitrogen and phosphorus and blooming of soybeans
  • Application of miR399 in promotion of absorption and utilization of nitrogen and phosphorus and blooming of soybeans

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Example 1 Construction of whole transgenic soybean lines overexpressing miR399e and miR399g

[0041] 1. Construction of miR399e and miR399g overexpression vectors

[0042] (1) Primer design

[0043] Download the full-length precursor sequences of miR399e and miR399g from the website, and design specific amplification primers, OE-miR399e (F( Sac I): AAgagctcCATGCAACGCAAAATAAAGG; R( Xba I ):AAtctagaAAATATGGCATATGAATGGT),

[0044] OE-miR399g(F( Sac I ): AAgagctcCCACCAACGCATATTATAACC; R( Xba I ): AAtctagaCATGAAGCAGTACTTTTAAGCC).

[0045] (2) Fragment amplification and loading pTF101.1

[0046] Using the cDNA of soybean YCO3-3 as a template, the corresponding fragments were amplified with specific primers of OE-miR399e and OE-miR399g respectively. The reaction system and reaction conditions are shown in Table 1 and Table 2 below.

[0047] Table 1 PCR reaction system

[0048]

[0049] Table 2 PCR reaction conditions

[0050]

[0051] (3) PCR amplification p...

Embodiment 2

[0070] Example 2 Analysis experiment of excess miR399e and miR399g

[0071] 1. Experimental method

[0072] (1) Seed disinfection

[0073] Mature, plump soybean seeds are arranged in a single layer in a Petri dish, placed in a desiccator, and the Petri dish is opened with the lid next to the Petri dish. Add 100 ml of sodium hypochlorite to a 250 ml beaker in a desiccator, then slowly add 4.2 ml of concentrated hydrochloric acid (HCl) along the wall of the beaker, immediately seal the desiccator, and let it stand for 10 hours. Remove excess chlorine after removal from desiccator.

[0074] (2) Germination

[0075] Put the sterilized seeds into the moist quartz sand to accelerate germination.

[0076] (3) Transplanting seedlings

[0077] One week later, when the two leaves are one heart, the normal seedlings with the same growth are selected and transplanted to the hydroponic system, and cultivated with 1 / 2 soybean nutrient solution in the greenhouse.

[0078] (4) Processin...

Embodiment 3

[0085] Example 3 Effects of excess miR399e and miR399g on the root system of transgenic soybean plants

[0086] 1. The root system is the main organ for plants to absorb phosphorus. The root surface area represents the contact area between the root system and the soil, and is an important indicator related to nutrient absorption.

[0087] We treated transgenic soybeans and wild-type soybeans overexpressing miR399e and miR399g for 40 days, and then collected samples to analyze root traits.

[0088] 2. Results

[0089] figure 2 The effect of excess miR399e and miR399g on the root system of transgenic soybean plants. The results showed that: under HPHN conditions, the total root length of the two OE-miR399e strains (#6, #7) was 1.85 and 1.72 times that of the wild type, respectively, and that of OE-miR399g-1 was 1.63 times that of the wild type. Under LPHN conditions, the total root lengths of two OE-miR399e lines (#6, #7) were 1.34 and 1.49 times that of the wild type, respe...

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Abstract

The invention discloses application of miR399 in the promotion of the absorption and utilization of nitrogen and phosphorus and blooming of soybeans. The miR399 has the functions of regulating and controlling the balance of phosphorus nutrient and blooming of the soybeans. By virtue of overexpression of miR399e/g, the total root length and root surface area can be increased, the contents of nitrogen and phosphorus of the soybeans can be increased, and therefore, the biomass is increased. According to the application, the capacities of absorbing and utilizing nitrogen and phosphorus of leguminous crops can be improved by virtue of a transgenic technique, and the application has important theoretical and practical significances for the adaption of low-phosphorous stress of the soybeans and the revelation of efficient synergy of nitrogen and phosphorus of the leguminous crops.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering. involves miR399 Application in promoting soybean nitrogen and phosphorus absorption and utilization and flowering. More specifically, soy miR399 The new function of promoting soybean root development, increasing nitrogen and phosphorus content and biomass, and promoting flowering, and the application of the gene in cultivating nitrogen and phosphorus synergistic high-efficiency plants and promoting flowering of leguminous crops. Background technique [0002] MicroRNA (miRNA) is a class of non-coding small molecule RNA with a length of 20-24 nucleotides, which exists in abundance in plants and can inhibit gene expression at the post-transcriptional level (Jones-Rhoades et al., 2006; Rogers et al. ., 2013). The biosynthesis process of miRNA in plants is complex (Kai et al., 2010). Previous studies have shown that miRNAs play a very important role in plant adaptation to nutrient str...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113A01H5/00
CPCC12N15/113C12N15/8243C12N2310/11
Inventor 王金祥陶平徐锋刘倩李波娣
Owner SOUTH CHINA AGRI UNIV
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