Streptomyces and application thereof

A Streptomyces, No. 1 technology, applied in the direction of bacteria, microorganisms, biochemical equipment and methods, etc., can solve the lack of ε-polylysine synthesis ability, low production capacity and level of ε-polylysine, and unsatisfactory issues such as increasing living standards

Active Publication Date: 2012-08-01
NANKAI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The production capacity and level of ε-polylysine in my country are relatively low, which cannot meet the needs of people's growing livi...

Method used

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  • Streptomyces and application thereof
  • Streptomyces and application thereof
  • Streptomyces and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1. Isolation and identification of Streptomyces sp.NK49

[0030] The experimental materials come from farmland soil in Shenyang, Liaoning Province. The specific steps are as follows:

[0031] Place the fresh soil samples taken in a cool and dry place for 3 days, then mix them evenly with 15% calcium carbonate, and then place them in a cool place to dry naturally for 8 days. Take 1g of the dried and sieved soil sample and 9ml of the diluent used to dilute the soil, oscillate and mix well, take the supernatant for gradient dilution, and apply the 100-fold and 1000-fold diluted supernatant to the solution added with 50mg / L K 2 Cr 2 o 7 The Gaoshi No. 1 medium plate was used for the isolation of ε-polylysine-producing strains; after 7 days of cultivation, the actinomycete colonies with dry and dense surface were selected, and the plate was streaked to contain no K 2 Cr 2 o 7 Purify colonies on the Gaoshi No. 1 medium plate, then pick independent colonies, inoc...

Embodiment 2

[0032] Embodiment 2, the shake flask method cultivation of starch hydrolysis sugar as single carbon source

[0033]Pick a ring of spores from Streptomyces sp.NK49 preserved on the slant of Benet and inoculate it into a 500ml Erlenmeyer flask containing 100ml of seed medium Gao's No. 1 medium, activate culture at 30°C, 180rpm, and activate for 24h The activated seed culture is inoculated into No. 1 Gao's culture medium with 10% inoculum size (adopting 500ml Erlenmeyer flask during shake flask fermentation, liquid capacity is 100ml) as seed liquid 30 ℃, 180rpm cultivates 24h; 15% of the inoculum size was inoculated in 10 bottles of starch hydrolysis sugar synthesis medium (500ml Erlenmeyer flask was used during the shake flask fermentation, and the filling volume was 100ml) to ferment at 30°C and cultivated at 180rpm for 80h; Centrifuge at 8000rpm for 20min, discard the bacteria, and collect the supernatant; use the D152 weak acid type cation exchange resin for product adsorptio...

Embodiment 3

[0034] Embodiment 3, with starch hydrolysis sugar and glycerol as the shaking flask method culture of mixed carbon source

[0035] (1) Pick a ring spore from the Streptomyces sp.NK49 preserved on the slope of Benet and inoculate it into a 500ml Erlenmeyer flask containing 100ml of seed medium Gaoshi No. 1 medium, and activate it at 30°C and 180rpm , Activation 24h; The activated seed culture is inoculated to Gao Shi No. 1 medium with 10% inoculum size (adopting 500ml Erlenmeyer flask during shake flask fermentation, and the filling capacity is 100ml) as seed liquid 30 ℃, 180rpm cultivates 24h; The seed solution was inoculated in 10 bottles of starch hydrolysis sugar and glycerol synthesis medium with 15% inoculum amount (500ml Erlenmeyer flask was used during the shake flask fermentation, and the liquid volume was 100ml) as the seed solution at 30°C, and cultivated at 180rpm for 96h; After the end, the culture was centrifuged at 8000rpm for 20min, the bacteria were discarded, ...

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Abstract

The invention discloses a streptomyces sp. NK-49 and an application thereof; and the application is a method for fermenting, cultivating, separating and purifying Epsilon-polylysine. The method comprises the following steps: firstly, picking a ring of spores from streptomyces sp. NK-49 which is stored on a Beinart inclined plane, and inoculating the spores serving as seeds in a Gao 1th culture medium for activated culture within 20 to 36 h; inoculating 5 to 15 percent of seed liquid to a synthetic medium taking amylolysis sugar or amylolysis sugar and glycerol as a carbon source, and shaking a flask for fermentation and culture for 48 to 96 h; after fermentation, conducting 6000 to 10000 rpm centrifugation on the fermentation liquid to remove thallus, collecting supernate, regulating the pH of the supernate to 8 by utilizing 5M NaOH solution, and filtering to removing deposited hetero proteins; utilizing weak acid type cation exchange resin to adsorb and remove Epsilon-polylysine molecules from the supernate of the hetero proteins, and washing with distilled water; eluting the adsorbed products by utilizing 0.1 to 0.7 mol/L dilute hydrochloric acid, and utilizing 5M NaOH solution to regulate the pH of eluant to a neutral value; and dialyzing the neutral eluant to remove micro molecular impurities, freezing, and drying to obtain Epsilon-polylysine solid powder. The streptomyces sp. NK-49 obtained through separation has the potential for industrial production of Epsilon-polylysine.

Description

Technical field: [0001] The present invention relates to Streptomyces NK49 and its use and method for producing amino acid homopolymer ε-polylysine with antibacterial activity. Background technique: [0002] ε-Poly-L-Lysine (ε-Poly-L-Lysine, ε-PL) is a homotype monomer polymerization formed by dehydration condensation of 25-30 L-lysine monomers through ε-amino and α-carboxyl groups polypeptide, its structural formula is: [0003] [0004] ε-polylysine is often produced by actinomycetes, has broad-spectrum antibacterial activity against Gram-negative bacteria, Gram-positive bacteria and fungi, and can withstand heat treatment during food processing, and can be used together with food raw materials Handle and process. ε-polylysine has been approved by the FDA as a food preservative. ε-polylysine has high safety, strong water solubility, and can be decomposed into the essential amino acid in the human body—lysine. It is a nutritional preservative. The production capacity...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P13/02C12R1/465
Inventor 宋存江谢晨庚耿伟涛王晓萌王淑芳赵若竹李毅翔杨超
Owner NANKAI UNIV
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