BVG90 _ 08615 gene deleted serratia marcescens engineering bacterium

A technology of Serratia marcescens and engineering bacteria, applied in the biological field, can solve the problems of hindering the industrialization process of microbial fermentation method and low yield.

Active Publication Date: 2020-09-29
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, there are still certain defects in the existing biological methods, among which the low yield is the most important defect hindering the industrialization of microbial fermentation methods

Method used

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  • BVG90 _ 08615 gene deleted serratia marcescens engineering bacterium
  • BVG90 _ 08615 gene deleted serratia marcescens engineering bacterium

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1: Construction of Serratia marcescens engineering bacteria JNB5-1ΔBVG90_08615

[0031] Specific steps are as follows:

[0032] With the genome of Serratia marcescens JNB5-1 as a template, respectively 08615-D-U-F / 08615-D-U-R (SEQ ID NO:3 and SEQ ID NO:4) and 08615-D-D-F / 08615-D-D-R (SEQ ID NO:5 and SEQ ID NO: 6) are primers, obtain DNA fragment BVG90_08615-U (SEQ ID NO: 7) and DNA fragment BVG90_08615-D (SEQ ID NO: 8) by PCR amplification; synthetic aacC3 resistance gene (SEQ ID NO: 9); DNA fragment BVG90_08615-U, DNA fragment BVG90_08615-D and aacC3 resistance gene were sequentially connected by overlap extension PCR to obtain DNA fragment BVG90_08615-AacC3; DNA fragment BVG90_08615-AacC3 was digested with KpnI linearized pUTmini The plasmids were ligated after homologous recombination to obtain the recombinant plasmid pUTmini-BVG90_08615; the recombinant plasmid pUTmini-BVG90_08615 was transformed into Escherichia coli (Escherichiacoli) S17-1 to obtain the t...

Embodiment 2

[0036] Embodiment 2: the production of prodigiosin

[0037] Specific steps are as follows:

[0038] Taking Serratia marcescens JNB5-1 as a control, the single bacterium colonies of Serratia marcescens engineering bacteria JNB5-1ΔBVG90_08615, JNB5-1ΔBVG90_03840 and JNB5-1ΔBVG90_24040 obtained in Example 1 were picked and inoculated into LB liquid medium ( Contains 50μg·mL -1 Apramycin and 50 μg·mL -1 Clindamycin), cultured with shaking at 37°C and 180rpm until the early logarithmic growth phase (OD 600 =0.6), to obtain a seed solution; the seed solution was inoculated into LB liquid medium with a 4% (v / v) inoculation amount, and fermented for 24 hours at 30° C. and 180 rpm to obtain a fermentation broth.

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Abstract

The invention discloses a BVG90 _ 08615 gene deleted serratia marcescens engineering bacterium, and belongs to the technical field of biology. The invention provides a serratia marcescens engineeringstrain JNB5-1 [delta] BVG90 _ 08615 capable of highly yielding prodigiosin. The serratia marcescens engineering strain JNB5-1 [delta] BVG90 _ 08615 is a serratia marcescens strain. The serratia marcescens engineering bacterium JNB5-1 delta BVG90 _ 08615 is obtained by knocking out a gene for coding a transcription regulation factor BVG90 _ 08615 in the serratia marcescens JNB5-1. The serratia marcescens engineering strain JNB5-1 delta BVG90 _ 08615 is inoculated into an LB liquid culture medium to be fermented for 24 h, and the yield of prodigiosin in fermentation liquor can reach 58.68 mg / L and is 1.15 times that of wild serratia marcescens JNB5-1.

Description

technical field [0001] The invention relates to Serratia marcescens engineering bacteria with BVG90_08615 gene deletion, and belongs to the field of biotechnology. Background technique [0002] Prodigiosin (PG) is a class of secondary metabolites produced by microorganisms. Studies have found that prodigiosin has a variety of biological activities, including: anti-bacteria, anti-dysentery, anti-tumor and immunosuppression, etc. Therefore, prodigiosin has great application value in fields such as pharmaceutical development. [0003] At present, the methods for producing prodigiosin mainly include chemical synthesis and microbial fermentation. Among them, the chemical synthesis method mainly obtains prodigiosin through tandem conjugate addition and high-temperature dehydrogenation. However, due to complex and difficult pathways and low yields, chemical synthesis is difficult to achieve large-scale industrial production. The principle of microbial fermentation is mainly to o...

Claims

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Application Information

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IPC IPC(8): C12N1/21C12P17/16C12R1/43
CPCC07K14/24C12P17/165
Inventor 饶志明潘学玮杨套伟尤甲甲易敢峰付维来徐美娟张显邵明龙
Owner JIANGNAN UNIV
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