Application of nKCBP protein in regulation and control of nitrogen fixation capability of leguminous plants

A leguminous plant and protein technology, applied in applications, plant peptides, plant products, etc., can solve the problems of destroying the ecological environment, green agricultural development, economic and resource waste, etc., to improve biological nitrogen fixation efficiency, reduce plant height and nitrogen fixation. Ability, the effect of significant application value

Active Publication Date: 2022-05-13
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The development of modern agriculture largely depends on the application of nitrogen fertilizer, which not only causes a huge waste of economy and resources, but also destroys the ecological environment and the development of green agriculture

Method used

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  • Application of nKCBP protein in regulation and control of nitrogen fixation capability of leguminous plants
  • Application of nKCBP protein in regulation and control of nitrogen fixation capability of leguminous plants
  • Application of nKCBP protein in regulation and control of nitrogen fixation capability of leguminous plants

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0087] Embodiment 1, evolution analysis of KCBP protein and tissue expression analysis of nKCBP protein

[0088] 1. Phylogenetic tree analysis of KCBP protein

[0089] The inventors of the present invention used the amino acid sequence of the KCBP protein from Arabidopsis to perform gene copy number and phylogenetic tree analysis on the KCBP proteins of 26 species (including 14 families of alfalfa).

[0090] see results figure 1 . The results showed that KCBP protein could amplify the genes of 7 species of rhizobia symbiotic nodulation in leguminous plants, increase the number of gene copies, and appear the phenomenon of gene duplication.

[0091] 2. Enrichment and expression of nKCBP protein in Medicago truncatula root nodules

[0092] Medicago truncatula has two homologous genes encoding KCBP protein, namely Medtr5g025100 and Medtr8g072430. Medtr8g072430 was named nKCBP gene (Nodulation-specific Kinesin-like Calmodulin-Binding Protein). The nucleotide sequence of the n...

Embodiment 2

[0109] Example 2, the acquisition of nKCBP gene knockout mutants and the application of nKCBP protein in reducing the nitrogen fixation ability of Medicago truncatula

[0110] 1. Construction of nKCBP gene knockout vector

[0111] The nKCBP gene knockout vector—recombinant plasmid pCambia1300-Cas9-nKCBP was constructed by CRISPR / Cas9 system. The target sequence is 5'-GTGATGGATATGACAGTGA-3' (that is, the 118th-137th position from the 5' end of SEQ ID No: 2). Vector construction methods refer to the following literature: Yan L, Wei S, Wu Y, Hu R, Li H, Yang W, Xie Q. High-Efficiency Genome Editing in Arabidopsis Using YAO Promoter-Driven CRISPR / Cas9 System.Mol.Plant.2015 8(12):1820-1823.

[0112] The construction process of the knockout vector pCambia1300-Cas9-nKCBP is as follows:

[0113] 1. Construction of sgRNA cassette

[0114] Insert the target sequence into the recognition site of the restriction endonuclease BsaI of the vector AtU6-26-sgRNA-SK to obtain an intermediat...

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Abstract

The invention discloses application of nKCBP protein in regulation and control of nitrogen fixation capacity of leguminous plants. The amino acid sequence of the nKCBP protein is as shown in SEQ ID No. 2. The inventor of the invention uses a CRISPR/Cas9 technology to edit the nKCBP gene at a fixed point and knock out the medicago truncatula nKCBP gene, so that the plant height and the nitrogen fixation capability are reduced. The invention is very important for researching the efficient nitrogen fixation mechanism of leguminous plants, and provides a new thought for improving the biological nitrogen fixation efficiency. The invention has important application value for plant breeding.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to the application of nKCBP protein in regulating the nitrogen fixation ability of leguminous plants. Background technique [0002] Legumes are important economic crops that play a role in agricultural production and economic development. Legumes and rhizobia can establish a mutually beneficial symbiotic relationship to achieve biological nitrogen fixation, that is, rhizobia transform molecular nitrogen in the atmosphere The process by which nitrogen is available for plant uptake. Symbiotic nitrogen fixation is the biological nitrogen fixation system with the highest nitrogen fixation capacity and the largest amount of nitrogen in nature. Effective use of this system is crucial to the sustainable development of agriculture and the nitrogen cycle in the biosphere. The development of modern agriculture largely depends on the application of nitrogen fertilizer, which not only ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/113C12N15/82A01H5/00A01H6/54
CPCC07K14/415C12N15/113C12N15/8218C12N15/8261C12N2310/20
Inventor 孔照胜张霞霞王琪黄弋戈吴菁霞
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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