Pseudoalteromonas polysaccharide degrading bacteria as well as culture method and application thereof

A kind of pseudoalteromonas, expanded culture technology, applied in the field of pseudoalteromonas polysaccharide degrading bacteria and its culture

Active Publication Date: 2019-04-09
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, rare polysaccharide degrading bacteria of the genus Pseudoalteromonas with glycosaminoglycan degrading ability

Method used

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  • Pseudoalteromonas polysaccharide degrading bacteria as well as culture method and application thereof
  • Pseudoalteromonas polysaccharide degrading bacteria as well as culture method and application thereof
  • Pseudoalteromonas polysaccharide degrading bacteria as well as culture method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Embodiment 1, separation and purification of coastal sediment microorganisms

[0069] Take the coastal sediments near the trestle bridge in Qingdao, take 1g sample, put it in the only carbon source medium of agarose, alginate or carrageenan with a volume of 100mL, under the conditions of the temperature of 28°C and the number of revolutions of 200 rpm, Cultivate until the solution is turbid; take 1 mL of the solution and add it to 9 mL of sterile water, and dilute to a concentration of 10 -1 、10 -2 、10 -3 、10 -4 、10 -5 5 concentration gradients. Spread the diluted bacterial suspension on the primary screening medium, make two parallels for each concentration, and culture at 28°C for 7 days. Pick out the single colony that can make the plate sink, and after 3 times of streaking on the plate to separate and purify, pick the single colony in the liquid medium for primary screening, culture at 28°C and 200 rpm for 3 days, take 1.6 mL of the culture and add 400 μL of gl...

Embodiment 2

[0075] Embodiment 2, the screening of polysaccharide degrading bacteria

[0076] The single clones of the strains obtained in Example 1 were inoculated into the sole carbon source liquid medium, cultured at 200 rpm and 30°C for 72 hours, and the turbidity of the bacterial solution and the change of the viscosity of the culture solution were observed. The enzyme-producing strains were selected according to the above two indicators. Pick the strains with large changes in the turbidity and viscosity of the bacteria in each unique carbon source medium, and pick them on the solid medium for primary screening and culture them, keep the strains and record them with numbers, such as Q01, Q02, Q03, ... etc.; after culturing a single colony for 24 hours, stain with modified Lugol's iodine solution;

[0077] The preparation method of the above-mentioned sole carbon source medium is:

[0078] Add polysaccharide substrates to the artificial seawater to a final concentration of 0.10% (w / v...

Embodiment 3

[0087] Example 3, Molecular identification of Pseudoalteromonas sp.Q02 strain based on 16S rRNA gene cloning

[0088] Genomic DNA of bacterial strain Q02 was prepared with a bacterial genome extraction kit (Tiangen Biochemical) and used as a template, amplified with universal primers (27f and 1492r) of bacterial 16S rRNA gene, and subjected to agarose gel electrophoresis, and then gel The recovery kit (Tiangen) purified the PCR amplification product, and after electrophoresis verification, it was connected to pEASY-Blunt Simple CloningVector and transformed into E.coli Trans1 T1 competent cells. After ampicillin resistance screening, positive clones were obtained. The obtained 16S rRNA gene sequencing was completed by Sangon Bioengineering (Shanghai) Co., Ltd. The sequence was compared with the 16S rRNA gene sequence of the standard strain collected by the National Center for Biological Information (NCBI) in the United States, and the phylogenetic tree was constructed using ME...

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Abstract

The invention discloses pseudoalteromonas polysaccharide degrading bacteria as well as a culture method and application thereof. The pseudoalteromonas sp. Q08 is preserved in China General Microbiological Culture Collection Center in Institute of Microbiology, Chinese Academy of Sciences No. 3, Courtyard 1, West Beichen Road, Chaoyang District, Beijing, with the preservation number being CGMCC No.16720. The strain can grow by taking microbe, alga, land plants and various types of animal-derived polysaccharide as an only carbon source, ensures that the species of produced polysaccharide degrading enzyme is rich, belongs to multifunctional polysaccharide degrading bacteria, and ensures that extracellular enzyme preparations prepared from the strain degrade microbe, alga, land plants and animal saccharides, and is obvious in degradation activity for sodium alginate and agarose from alga and HA, CSC and CSE from animals.

Description

technical field [0001] The invention relates to a polysaccharide-degrading bacterium belonging to the genus Pseudoalteromonas and its cultivation method and application, belonging to the field of biotechnology. Background technique [0002] Polysaccharides are polymers composed of repeating single or different sugar units connected by glycosidic bonds. According to different biological sources, common microbial polysaccharides include peptidoglycan, xanthan gum, etc., seaweed polysaccharides such as agarose, alginate, carrageenan, etc., higher plant polysaccharides such as cellulose, mannan, xylan and starch, etc. And animal polysaccharides such as glycosaminoglycans, chitin, etc. [1] . These polysaccharides are important components of organisms, some are energy substances, and some are information molecules involved in cell recognition [2] . Among them, glycosaminoglycans or oligosaccharides, including hyaluronic acid, chondroitin sulfate / dermatan sulfate, heparin / hepar...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P19/14C12P19/04C12P19/00C12R1/01
CPCC12N1/20C12P19/00C12P19/04C12P19/14C12N1/205C12R2001/01
Inventor 韩文君王延辉李俊鸽单宁宁胡玮冯璋王一兵李福川古静燕
Owner SHANDONG UNIV
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