Ammonium-resistant nitrogen-fixing microorganism of which gene glnA is lost and constructing method and application thereof

A technology of nitrogen-fixing microorganisms and gene deletion, applied in the field of microorganisms, can solve the problems of low nitrogen fixation efficiency and high nitrogen fixation efficiency, achieve breakthrough inhibition, improve nitrogenase activity, and have wide application prospects

Active Publication Date: 2018-05-18
HUBEI FORBON TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In other words, nitrogen-fixing bacteria are highly efficient in nitrogen fixation in poor soils, but inefficient in fertile soils

Method used

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  • Ammonium-resistant nitrogen-fixing microorganism of which gene glnA is lost and constructing method and application thereof
  • Ammonium-resistant nitrogen-fixing microorganism of which gene glnA is lost and constructing method and application thereof
  • Ammonium-resistant nitrogen-fixing microorganism of which gene glnA is lost and constructing method and application thereof

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Experimental program
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Effect test

Embodiment 1

[0029] 1. Screening and construction of glnA mutant strains

[0030] The product of glnA gene is glutamine synthetase (GS). There are two glnA genes in Paenibacillus polymyxa, one of which is connected with glnR gene to form a transcription unit: glnRA, and the other glnA1 gene exists alone.

[0031] The present invention only deletes the glnA gene in the glnRA transcription unit.

[0032] Use the primers up(glnRA)-F / up(glnA)-R in Table 1 to amplify the upstream homology arm (1001bp) of glnA, and primer down(glnA)-F / down(glnA)-R to amplify the downstream homology arm (511bp), the vector pRN5101 was treated with BamHI / SalI restriction endonuclease, and then the above three DNA fragments were assembled using Gibson assembly master mix (New England Biolabs), and the assembled recombinant plasmid was transformed into Paenibacillus Azopolymyxa polymyxa, use the erythromycin resistance carried by the vector plasmid pRN5101 to screen single-exchange strains. After multiple generations of ...

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Abstract

The invention relates to the field of microorganisms, and particularly discloses an ammonium-resistant nitrogen-fixing microorganism of which the gene glnA is lost and a constructing method and application thereof. The ammonium-resistant nitrogen-fixing microorganism of which the gene glnA is lost can be subjected to biological nitrogen fixation under both an ammonium-free condition and an ammonium condition, the activity of dinitrogenase of paenibacillus microorganisms under a high-ammonium condition is improved, the inhibiting effect of biological nitrogen fixing under a high-ammonium condition is broken through, and the ammonium-resistant nitrogen-fixing microorganism has a wide application prospect in agricultural production.

Description

Technical field [0001] The present invention relates to the field of microorganisms, in particular to a glnA gene-deficient ammonium nitrogen-fixing microorganism, and a construction method and application thereof. Background technique [0002] Nitrogen is the largest element necessary for plant production. Chemical nitrogen fertilizers play a very important role in ensuring my country's food production, vegetable planting and fruit tree cultivation. However, long-term excessive partial application of chemical nitrogen fertilizers has led to soil acidification and salinization, and decreased soil microbial activity, which has become an important constraint on the sustainable development of agriculture. [0003] Biological nitrogen fixation is a small number of bacteria that reduce nitrogen in the air to ammonium at normal temperature and pressure. The ammonium formed by nitrogen fixation is absorbed and utilized by plants. However, biological nitrogen fixation is an energy-intens...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/21C12R1/01
CPCC12N9/93C12Y603/01002
Inventor 陈三凤赵喜云王天舒
Owner HUBEI FORBON TECH
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