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Lysinibacillus fusiformis ZJB-17006 and application thereof

A ZJB-17006, Bacillus lysine technology, applied in bacteria, microorganisms, biochemical equipment and methods, etc., can solve the problems of waste of raw materials, unconverted D-glufosinate, low product yield and e.e. value, etc. , to achieve the effect of mild catalytic reaction conditions, important application prospects, and easy cultivation, collection and application.

Active Publication Date: 2019-01-11
ZHEJIANG UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

3) Asymmetric synthesis method, asymmetric catalytic hydrogenation - large amount of catalyst and expensive trimethylsilyl cyanide; asymmetric Strecker reaction - use of highly toxic cyanide; asymmetric Michael addition - large amount of catalyst, And the product yield and e.e. value are low
4) The racemate resolution method, the highest yield of this method is 86%, the highest e.e. value is 99%, but after the resolution, D-glufosinate-ammonium is not converted, wasting raw materials

Method used

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  • Lysinibacillus fusiformis ZJB-17006 and application thereof
  • Lysinibacillus fusiformis ZJB-17006 and application thereof
  • Lysinibacillus fusiformis ZJB-17006 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1: Screening of spindle-shaped lysine bacillus (Lysinibacillus fusiformis) ZJB-17006

[0027] 1. Primary screening

[0028] The present invention takes soil samples from all over the country, and takes 80 parts of soil samples altogether. The specific method of screening: Weigh 1g of soil sample and place it in 10mL of 0.85% physiological saline, shake it and let it stand still, take the supernatant into the enrichment medium, and cultivate it at 30°C and 150r / min for 2-3 days with shaking . Take 1mL of the enrichment solution and add it to 50mL of fresh enrichment medium, and repeat this process 3 times before separation and purification.

[0029] Bromothymol blue filter paper was selected as the indicator filter paper to detect the colony capable of degrading the substrate. The principle is: after the colony containing the target enzyme degrades the substrate, acidic by-products (phenylacetic acid, acetic acid, formic acid, benzoic acid) will be produced, t...

Embodiment 2

[0045] Embodiment 2: bacterial strain ZJB-17006 identification

[0046] 1. Morphological identification:

[0047] The bacterial strain ZJB-17006 screened in Example 1 of the present invention forms a round or nearly round, soft texture, smooth surface, flat, neat edges, and shiny light yellow colony after being cultivated on a solid medium for 24 hours at 37°C , 2-4mm in diameter. Observation by Gram staining: purple long rod with spores. Composition of solid medium: sodium chloride 10g / L, peptone 10g / L, yeast powder 5g / L, agar 20g / L, solvent is deionized water.

[0048] 2. Physiological and biochemical identification:

[0049] Using the Biolog (GENⅢ) automatic microbial identification system, 94 kinds of phenotypic tests were carried out on the strain ZJB-17006, including 71 kinds of carbon source utilization detection and 23 kinds of chemical sensitivity detection: inoculate the strain on a specific plate medium, 33 ℃ Cultivate at constant temperature for 2 days, wash th...

Embodiment 3

[0059] Embodiment 3: the preparation of wet thalline

[0060] (1) Incline cultivation:

[0061] Spindle-shaped lysinibacillus ZJB-17006 was inoculated into the slant medium, and cultured at 30°C for 48 hours to obtain slant bacteria;

[0062] The final concentration of the slant culture medium is: casein peptone 17g / L, Na 2 HPO 4 3.0g / L, K 2 HPO 4 1.5g / L, soybean peptone 3g / L, glucose 2.5g / L, NaCl 5g / L, agar 20g / L, solvent is deionized water, pH value is 7.0.

[0063] (2) Seed cultivation

[0064] Pick an inoculation loop thalline from the slant thallus and inoculate it into the seed medium, and cultivate it at 30°C for 24 hours to obtain the seed liquid; the final concentration of the seed medium consists of: casein peptone 17g / L, Na 2 HPO 4 3.0g / L, K 2 HPO 4 1.5g / L, soybean peptone 3g / L, glucose 2.5g / L, NaCl 5g / L, solvent is deionized water, pH value is 7.0.

[0065] (3) Fermentation culture

[0066] The seed solution was inoculated into the fermentation mediu...

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Abstract

The invention relates to a Lysinibacillus fusiformis ZJB-17006 and application thereof. The preparation of L-amino acids by catalyzing N-phenylacetyl-DL-amino acid has an enantioselectivity value reaching 99%, and the preparation of 2-amino-4-[hydroxy(methyl)phosphoryl]-L-butyric acid by catalyzing 2-N-phenylacetyl-4-[hydroxy(methyl)phosphoryl]-DL-butyric acid has an enantioselectivity value reaching 99%.

Description

(1) Technical field [0001] The present invention relates to a bacterial strain producing amidohydrolase - Lysinibacillus fusiformis ZJB-17006, and its N- Application of phenylacetyl substituents in the preparation of chiral amino acid derivatives. (2) Background technology [0002] The chemical name of L-glufosinate-ammonium is: 4-[hydroxyl (methyl)phosphono]-L-homoalanine, which is a structural analog of L-glutamic acid and can inhibit glutamine synthetase (GS ) activity, the accumulation of ammonia in plants, the accumulation of high-concentration ammonia gas blocks the photorespiration of plants, the destruction of chloroplast structure and the vesicleization of matrix, and the synthesis of amino acids is blocked, resulting in cell membrane damage and cell death, thus killing weeds . It has a broad spectrum and is the second most herbicide-tolerant genetically modified crop in the world. [0003] Currently L-glufosinate-ammonium synthesis methods include chemical synth...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P13/00C12P13/06C12P13/14C12P13/22C12P13/08C12P13/20C12P13/12C12R1/01
CPCC12N1/20C12P13/001C12P13/06C12P13/08C12P13/12C12P13/14C12P13/20C12P13/222C12P13/225C12N1/205C12R2001/01
Inventor 柳志强郑裕国康雪梅张晓健金利群
Owner ZHEJIANG UNIV OF TECH
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