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Mutant strain capable of realizing high-efficiency production of low-molecular pulullan and application thereof

A pullulan polysaccharide and low-molecular-weight technology is applied in the field of mutagenic strains to achieve the effects of improving yield, mild production method conditions, and reducing production costs

Active Publication Date: 2018-03-06
QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] The present invention aims at the breeding and screening method for high-efficiency production of low-molecular pullulan obtained by mutagenesis in the existing pullulan production industry and the process of producing low-molecular pullulan using the method. The present invention provides a A mutagenic strain for efficient production of low-molecular-weight pullulan and its application, specifically relating to a low-molecular-weight pullulan obtained through directed evolution and mutagenesis screening using a commercial strain of Aureobasidium pullulans (A.pullulans CGMCC 3.933) The mutagenic strain of Aureobasidium pullulans UVMU6-1 (Aureobasidium pullulans), the strain preservation number is CGMCC No.13178 and the method for efficiently fermenting and producing pullulan using the mutagenic strain

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  • Mutant strain capable of realizing high-efficiency production of low-molecular pulullan and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0040] Example 1: Mutation and screening of high-efficiency low-molecular-weight pullulan strains

[0041] Get 10ml of Aureobasidium pullulans (A.pullulans) CGMCC 3.933 bacterial suspension (unit bacterium concentration 10 5 ~10 7 CFU / mL) in a petri dish with a diameter of 9 cm, irradiated with ultraviolet light under magnetic stirring. The power of the ultraviolet lamp is 10W-15W, and the irradiation distance is 30cm. Before the irradiation, turn on the ultraviolet lamp for 5min-20min to stabilize it. Cover the lid of the dish, turn off the ultraviolet light, take 100 μL of the bacterial solution and add it to the solid medium containing YPD (containing 0.03-0.07% Tritribe blue) to smear a plate, place it at 30°C, and keep it upside down for 2 days at a constant temperature. The depth and surface texture are the screening indicators, and the strains with dark color, wet colony surface and high ratio of colony diameter to thickness are screened out and numbered. Then it was...

Embodiment 2

[0045] Example 2: Determination of low molecular weight pullulan

[0046] Both the fermentation product and pullulan were dissolved in deionized water at a ratio of 1g / 10mL to obtain a sample solution and pullulan standard solution; maltotriose and glucose were dissolved in deionized water at a ratio of 10mg / mL; Layer system: Acetonitrile and distilled water are mixed evenly according to the volume ratio of n-butanol: acetic acid: distilled water = 8:3:2, and placed in a stoppered glass bottle for later use. Color developer: Concentrated sulfuric acid and absolute ethanol are mixed evenly according to the ratio of V concentrated sulfuric acid: V absolute ethanol = 1:6, and put into a watering can for later use.

[0047] Dilute the pullulan standard solution, pullulan polysaccharide hydrolyzate, sample solution, and sample enzymatic hydrolyzate by 100 times respectively, according to the standard solution of pullulan polysaccharide, pullulan polysaccharide hydrolyzate, maltotri...

Embodiment 3

[0047] Dilute the pullulan standard solution, pullulan polysaccharide hydrolyzate, sample solution, and sample enzymatic hydrolyzate by 100 times respectively, according to the standard solution of pullulan polysaccharide, pullulan polysaccharide hydrolyzate, maltotriose, and sample enzymatic hydrolyzate Spot the sample on the silica gel chromatographic plate in order of , sample solution, and glucose, and perform chromatography in the developing solution. After the liquid chromatography reaches the top of the silica gel plate, take it out and dry it, and repeat the chromatography 2-3 times. Spray the chromogenic agent onto the silica gel plate after chromatography, blow it dry, heat it on an alcohol lamp to develop color, and observe the experimental phenomenon. The separated obtained sample was hydrolyzed into maltotriose the same as the standard product, and the results showed that the obtained product was pullulan ( figure 1 ). Example 3: Efficient production of low-molec...

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Abstract

The invention discloses a mutant strain capable of realizing high-efficiency production of low-molecular pulullan and application thereof, belonging to the technical field of mutagenesis of strains. The Aureobasidium pullulans UVMU6-1 capable of realizing high-efficiency production of low-molecular pulullan in the invention has an access number of CGMCC No.13178. The invention also provides application of the Aureobasidium pullulans UVMU6-1 to production of low-molecular pulullan. The strain provided by the invention has high pulullan production capability and can effectively improve the yieldof pulullan and reduce production cost; since the strain has low viscosity during fermentation, the concentration of produced pulullan in fermentation broth is high, and fermentation cost is effectively reduced; and a production method for producing pulullan via the strain is mild in conditions and friendly to environment, and the produced pulullan is green, natural, etc.

Description

technical field [0001] The invention relates to a mutagenic strain for efficiently producing low-molecular-weight pullulan and an application thereof, belonging to the technical field of strain mutagenesis. Background technique [0002] Pullulan, also called pullulan, pullulan, pullulan or pullulan, is an exopolysaccharide produced by the fermentation of Aureobasidium pullulans. It is a tasteless and odorless white powder. The chemical composition of pullulan polysaccharide is mainly polymaltotriose linked by α~1,4~glucosidic bonds, with a molecular weight of 4.8×10 4 ~2.2×10 6 (The average molecular weight of commercial pullulan polysaccharide is 2×10 5 , composed of about 480 maltotriose). The strain of pullulan biosynthesis is mainly Pullulan yeast (Aureobasidium pullulans, A. pullulans), and the production raw materials include glucose, sucrose, starch, corn and other related biomass. my country approved pullulan as a new food additive in 2006. In Japan, pullulan is...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/14C12P19/04C12R1/645
CPCC12P19/04C12N1/145C12R2001/645
Inventor 张海波咸漠陈国强王纪明赵宏伟刘会洲刘炜
Owner QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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