Engineering bacteria based on nitrate reductase and implementation method of engineering bacteria

Abstract

The invention relates to engineering bacteria based on nitrate reductase and an implementation method of the engineering bacteria. The method includes the steps of firstly, conducting PCR amplification on primers containing enzyme cutting sites with the streptomyces griseorubens genome DNA as a template to sequentially obtain the nucleotide sequence of encoding nitrate reductase catalysis subunits and the nucleotide sequence of electron transfer subunits; secondly, sequentially connecting the gene sequence obtained through amplification to the co-expression vector pETDuet-1; thirdly, obtaining the recombination co-expression vector pETDuet-NB-NA; fourthly, converting the nitrate reductase expression vector into escherichia coli expression strains. In order to overcome the defects that nitrate reductase is generally endoenzyme in vivo and the expression quantity is low, the application range and the effect are seriously limited, a large amount of nitrate reductase can be expressed and synthesized in vitro through the genetic engineering means. The engineering bacteria and the method have great significance in rapid repair of secondary salinization soil and even the achievement of precision agriculture.

Description

technical field [0001] The invention relates to a gene in the technical field of biogenetic engineering and its engineering strain, in particular to a nitrate reductase-based engineering bacterium of Streptomyces grisea and its realization method. Background technique [0002] With the development of facility agriculture and the continuous expansion of greenhouse cultivation areas, the use of nitrogen fertilizers has increased significantly, greatly exceeding the demand of plants. Because the excess nitrate in the soil was not absorbed and transformed by plants in time, a large amount of salt ions accumulated in the soil, and eventually secondary salinization of the facility carrier soil was formed. [0003] Studies have shown that HCO removal in protected cultivation secondary salinized soil 3 － Outside, Na + 、K + , Ca 2+ , Mg 2+ , Cl － , SO 4 2－ , NO 3 － have different degrees of accumulation. Studies have shown that the cation composition in this type of soil,...

AI Technical Summary

Problems solved by technology

Although there are many examples of successful application of these methods, there are also many shortcomings: irrigation and salt washing will wash nitrate nitrogen into the ground, which not only causes the loss of soil nitrogen, but also pollutes groundwater; High, easy to produce secondary pollution; the semi-decomposed organic fertilizer method has the disadvantages of slow fertilizer efficiency and inconvenient use

However, the expression level of nitrate reductase in microorganisms is generally very low, and most of them are expressed intracellularly, so the scope and effect of application are very limited and cannot meet the needs of repairing the environment

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