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Application of Peanut ahfrdl1 Gene in Improving Plant Resistance to Aluminum Toxic Stress

A peanut and genetic technology, applied in the field of genetic engineering, to achieve the effect of improving yield and quality and having broad application prospects

Active Publication Date: 2017-10-27
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in peanut, the MATE gene related to aluminum stress resistance has not yet been reported.

Method used

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  • Application of Peanut ahfrdl1 Gene in Improving Plant Resistance to Aluminum Toxic Stress
  • Application of Peanut ahfrdl1 Gene in Improving Plant Resistance to Aluminum Toxic Stress
  • Application of Peanut ahfrdl1 Gene in Improving Plant Resistance to Aluminum Toxic Stress

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Cloning of Example 1 Peanut AhFRDL1 Gene

[0049] (1) Hydroponic cultivation of peanuts

[0050] Peanut seeds Luhua 14 (purchased from the Chinese Academy of Agricultural Sciences) were sterilized with 10% hydrogen peroxide for 30 minutes, cleaned, placed in saturated calcium sulfate solution for about 8 hours, and ventilated to avoid light. Then place them in a tray covered with absorbent paper, pour an appropriate amount of water, and place them in an artificial cultivation room in the dark. The culture room was set at 30°C for 14 hours with light and 22°C for 10 hours with darkness. About 1 day after the peanuts sprouted small buds, they were transferred to quartz sand for cultivation, and after 2 leaves grew, they were transferred to hydroponic nutrient solution for cultivation. The formula of hydroponic nutrient solution is as follows: KH 2 PO 4 (0.5mM), K 2 SO 4 (0.75mM), KCl (0.1mM), MgSO 4· 7H 2 O (0.65mM), CaSO 4· 2H 2 O(2mM), H 3 BO 3 (1μM), MnSO ...

Embodiment 2

[0060] The construction of embodiment 2 recombinant expression vector

[0061] 1. Build the carrier

[0062](1) Add restriction sites XmaI and BamHI through primers, and use the vector pMD20-T-AhFRDL1 as a template to perform PCR;

[0063] The primer sequences are:

[0064] 35s-AhFRD1-F: 5'-ACGCCCCGGGATGGCTGAGAAGCAG-3' (SEQ ID NO.5);

[0065] 35s-AhFRD1-R: 5'-CGGGATCCTTTTCTCATAGGAATTCCCAAG-3' (SEQ ID NO.6);

[0066] (2) Cloning the PCR product into the pGEM-T vector, transforming DH5α competent cells, selecting positive clones, culturing the bacteria and extracting the plasmid to obtain the vector pGEM-T-AhFRDL1;

[0067] (3) The vector pGEM-T-AhFRDL1 and pBAR1 vectors were double-digested with XmaI and BamHI at the same time, and the target fragment of the AhFRDL1 gene and the pBAR1 vector fragment were recovered respectively, and then the target fragment of the AhFRDL1 gene and the digested pBAR1 were ligated with T4 ligase Vector fragment, constructing a 35S strong prom...

Embodiment 3

[0080] Example 3 Functional complementation test of peanut AhFRDL1 gene to Arabidopsis single mutant AtALMT1-Ko, AtMATE-Ko and double mutant Atdouble-Ko

[0081] 1.1 Acquisition of Arabidopsis plants transfected with AhFRDL1 gene

[0082] The recombinant expression vector pBAR1-AhFRDL1 obtained in Example 2 was transformed into three Arabidopsis mutants lacking the function of aluminum resistance through transgenic technology: (1) Arabidopsis aluminum-resistant gene AtALMT1 (malic acid transport carrier) knockout Single mutant AtALMT1-Ko; (2) single mutant AtMATE-Ko knocked out of Arabidopsis aluminum tolerance gene AtMATE (citrate transport carrier); (3) Arabidopsis double mutant knocked out of both AtALMT1 and AtMATE Mutant Atdouble-Ko.

[0083] The specific operation steps are as follows: Soak the Arabidopsis seeds with 1% agarose, put the seeds into the nutrient soil mixed with vermiculite with a pipette gun (the ratio of vermiculite to nutrient soil is 1:1, and put them ...

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Abstract

The invention provides the application of peanut citrate transporter gene AhFRDL1 in improving plant resistance to aluminum toxicity stress. The application is to transfer the peanut citrate transporter gene AhFRDL1 or a recombinant expression vector containing the peanut citrate transporter gene AhFRDL1 into plant cells, screen transgenic plants, and make the transgenic plants express the peanut citrate transporter, thereby Improve plant resistance to aluminum toxicity stress. The present invention proves for the first time that the peanut citrate transporter gene AhFRDL1 is a gene that regulates the secretion of citric acid to the plant, and participates in the biological process of plants resisting aluminum toxicity stress. The peanut citrate transporter gene AhFRDL1 is used to improve the resistance of plants Under the stress of aluminum toxicity, it has long-term practical significance to further cultivate the main crops resistant to aluminum toxicity on acidic soil.

Description

technical field [0001] The invention belongs to the field of genetic engineering, and particularly relates to the application of a peanut citrate transporter gene AhFRDL1 in improving plant resistance to aluminum toxicity stress. Background technique [0002] Aluminum is the most abundant metal element in the earth's crust, usually in the form of insoluble silicate or alumina, which is not harmful to plants, but under acidic conditions (pH<5), soluble aluminum (mainly Al 3+ ) will be toxic to most plants. The acidic soil in my country accounts for about 21% of the total land area of ​​the country. Aluminum is not only the main source of soil acidity on acidic soil, but also because the exchange volume of aluminum accounts for 20-80% of the soil cation exchange volume. , leading to the easy leaching of cations in the soil, resulting in the lack of nutrients such as P, K, Ca, Mg, B, Mo, etc. Therefore, aluminum toxicity has become an important limiting factor restricting the...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/84A01H5/00
Inventor 左元梅邱巍纪春巧熊宏春郭笑彤
Owner CHINA AGRI UNIV
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