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Pseudomonas chlororaphis QOHPHZ-8 for producing 1-hydroxyphenazine and application

A technology of QOHPHZ-8 and QOHPHZ-1, applied in the field of microbial engineering strains, can solve the problems of low fermentation units of biocontrol Pseudomonas, and achieve the effects of low pathogenicity and increased yield

Active Publication Date: 2020-12-18
QILU UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At the same time, biopesticides are also facing bottlenecks in development, such as the low fermentation units of biocontrol Pseudomonas, etc.

Method used

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  • Pseudomonas chlororaphis QOHPHZ-8 for producing 1-hydroxyphenazine and application
  • Pseudomonas chlororaphis QOHPHZ-8 for producing 1-hydroxyphenazine and application
  • Pseudomonas chlororaphis QOHPHZ-8 for producing 1-hydroxyphenazine and application

Examples

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Embodiment 1

[0051] In the present embodiment, a method for preparing Pseudomonas chlororaphis QOHPHZ-1 based on Pseudomonas chlororaphis Qlu-1 is provided, and the steps of the preparation method are as follows:

[0052] 1. Inoculate Pseudomonas chloropinus Qlu-1 into KB(A+) medium, culture overnight at 30°C with shaking at 180rpm, use the genome extraction kit to extract the genome of Qlu-1, and store it at -20°C for later use.

[0053] 2. Search the phzO gene and its upstream and downstream sequences in the sequenced Qlu-1 genome data, use the Qlu-1 strain genome as a template, and use phzO-F1 / phzO-R1, phzO-F2 / phzO-R2 as primers respectively Amplify the upstream fragment phzO-U and the downstream fragment phzO-D of the phzO gene; use phzO-U and phzO-D as templates, and phzO-F1 / phzO-R2 as templates to amplify the phzO upstream and downstream fusion fragment phzO-UD. The fusion fragment phzO-UD was ligated with the knockout plasmid pk18moBsacB to construct the recombinant plasmid pk18-phz...

Embodiment 2

[0058] In the present embodiment, a method for preparing Pseudomonas chlororaphis (Pseudomonas chlorophis) based on Pseudomonas chlororaphis QOHPHZ-1 described in Example 1 is provided.

[0059] The method of QOHPHZ-8, described preparation method comprises the following steps:

[0060] On the basis of QOHPHZ-1, genes phzC, aroB, aroD, aroE related to the leading pathway of phenazine synthesis and central metabolism related genes tktA, ppsA were amplified respectively. And the Bglbric module plasmid pBbB8k-GFP was used as the intermediate particle to construct a single module plasmid, and the gene was integrated into a tandem gene pBbB8K-tktA-ppsA-phzC-aroB-aroD-aroE by using the module principle.

[0061] The tandem gene was excised from pBbB8K-tktA-ppsA-phzC-aroB-aroD-aroE, ligated into the integration plasmid pUC18-mini-Tn7T-Gm, and integrated through the pUC18-mini-Tn7T-Gm plasmid with the help of the helper plasmid pTNS2 Enter the QOHPHZ-1 genome to obtain Pseudomonas ch...

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Abstract

The invention particularly relates to pseudomonas chlororaphis QOHPHZ8 for producing 1-hydroxyphenazine and application. A pseudomonas chlororaphis Qlu-1 strain is obtained by screening and has a typical gene cluster synthesized by phenazine compounds, and the strain is natural and cannot synthesize and secrete 1-OH-PHZ. Firstly, a phzO gene is replaced by phzS to prepare QOHPHZ-1, and 1-OH-PHZ accumulation is obtained; in order to further obtain a 1-hydroxyphenazine high-yield strain, subsequent optimization work is carried out on a QOHPHZ-1 strain, a QOHPHZ-8 strain is prepared, the yield of1-OH-PHZ in unit time is increased by 28 times or more, and the strain is expected to be an engineering strain to be applied to the fields of agricultural production and algae prevention and control.

Description

technical field [0001] The invention belongs to the technical field of microbial engineering strains, in particular to a Pseudomonas chlororaphis QOHPHZ-8 producing 1-hydroxy-phenazine (1-OH-PHZ), and the strain is used as an engineering strain application. Background technique [0002] The information disclosed in this background section is only intended to increase the understanding of the general background of the present invention, and is not necessarily taken as an acknowledgment or any form of suggestion that the information constitutes the prior art already known to those skilled in the art. [0003] Pesticides are important materials for agricultural production and are also of great significance for maintaining the agricultural ecological environment. Compared with chemical pesticides, biological pesticides have the characteristics of safety to non-target organisms, low toxicity and side effects, and good environmental compatibility, and have good development prospe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/21C12P17/12C05F11/08C07D241/44A01N43/60A01N63/27A01P13/00C12R1/385
CPCC12N9/0073C12P17/12C05F11/08C07D241/44A01N43/60A01N63/27
Inventor 刘开泉李玲王瑞明王腾飞李丕武
Owner QILU UNIV OF TECH