A kind of ammonium-resistant nitrogen-fixing microorganism with glna gene deletion and its construction method and application

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: 2021-04-27
HUBEI FORBON TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

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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  • A kind of ammonium-resistant nitrogen-fixing microorganism with glna gene deletion and its construction method and application
  • A kind of ammonium-resistant nitrogen-fixing microorganism with glna gene deletion and its construction method and application
  • A kind of ammonium-resistant nitrogen-fixing microorganism with glna gene deletion and its construction method and application

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Experimental program
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Embodiment 1

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

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

[0031] In the present invention, only the glnA gene in the glnRA transcription unit is deleted.

[0032] Use the primers in Table 1 up(glnRA)-F / up(glnA)-R to amplify the upstream homology arm (1001bp) of glnA, and the primers 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 nitrogen-fixing polymyxa polymyxa, using the erythromycin resistance carried by the vector plasmid pRN5101 to screen single-crosso...

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Abstract

The invention relates to the field of microorganisms, and specifically discloses an ammonium-resistant nitrogen-fixing microorganism with glnA gene deletion, a construction method and application thereof. The ammonium-resistant nitrogen-fixing microorganisms lacking the glnA gene can perform biological nitrogen fixation under ammonium-free and ammonium-containing conditions, improve the nitrogenase activity of Paenibacillus microorganisms under high ammonium conditions, and break through the limitations of high ammonium conditions on biological nitrogen fixation. Inhibition has broad application prospects in agricultural production.

Description

technical field [0001] The invention relates to the field of microorganisms, in particular to an ammonium-resistant nitrogen-fixing microorganism with glnA gene deletion and its construction method and application. Background technique [0002] Nitrogen is the largest element necessary for plant production. Chemical nitrogen fertilizer plays a very important role in ensuring my country's grain production, vegetable planting and fruit tree cultivation. However, long-term excessive application of chemical nitrogen fertilizers leads to soil acidification and salinization, and the decline of soil microbial activity has become an important factor restricting the sustainable development of agriculture. [0003] Biological nitrogen fixation is a process in which a small number of bacteria reduce nitrogen in the air to ammonium at normal temperature and pressure, and the ammonium formed by nitrogen fixation is absorbed and utilized by plants. However, biological nitrogen fixation ...

Claims

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Application Information

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Patent Type & AuthorityPatents(China)
IPC IPC(8): C12N1/21C12R1/12C12R1/01
CPCC12N9/93C12Y603/01002
Inventor陈三凤赵喜云王天舒
OwnerHUBEI FORBON TECH