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A Corynebacterium glutamicum engineering strain that biosynthesizes rare sugars and its construction method and application

A technology of Corynebacterium glutamicum and engineering strains, applied in the fermentative production of rare ketose and deoxyketose, the field of engineering strains of Corynebacterium glutamicum and its construction, can solve problems such as no relevant reports

Active Publication Date: 2016-03-23
天津怡和生物科技有限责任公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there are no relevant reports at home and abroad on the patents for the synthesis of D-sorbose, D-psicose and L-fructose by constructing recombinant strains

Method used

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  • A Corynebacterium glutamicum engineering strain that biosynthesizes rare sugars and its construction method and application
  • A Corynebacterium glutamicum engineering strain that biosynthesizes rare sugars and its construction method and application
  • A Corynebacterium glutamicum engineering strain that biosynthesizes rare sugars and its construction method and application

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Experimental program
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Embodiment 1

[0051] Embodiment 1, construction Corynebacterium glutamicum (Corynebacterium glutamicum) engineering strain SY10

[0052] The construction of Corynebacterium glutamicum (Corynebacterium glutamicum) engineering strain SY10 comprises the following steps:

[0053] 1. Knock out the triose phosphate isomerase gene in Corynebacterium glutamicum ATCC13032 to obtain a recombinant Corynebacterium glutamicum without triose phosphate isomerase activity, named SY6.

[0054] 1. Construction of triose phosphate isomerase gene (tpi) knockout vector pK18tip

[0055] 1.1 According to the nucleosides of the upstream region tpi' (964bp, sequence 1 in the sequence listing) and the downstream region tpi'' (861bp, sequence 2 in the sequence listing) of the Genbank Corynebacterium glutamicum ATCC13032tpi gene (Genbank number: 1019554) Acid sequence design primer 1 and primer 2, primer 3 and primer 4, primer 5 and primer 6, wherein primer 5 and primer 6 have XbaI and HindIII restriction sites, prim...

Embodiment 2

[0098] Embodiment 2, the application of Corynebacterium glutamicum (Corynebacterium glutamicum) engineering strain SY10 in producing rare ketose

[0099] 1. Application of Corynebacterium glutamicum engineering strain SY10 in the production of D-allose and D-sorbose

[0100] 1. Cultivation and induction of Corynebacterium glutamicum engineering strain SY10

[0101] Select CGXII medium (recipe: (NH 4 ) 2 SO 4 (5g / L),urea(5g / L),KH 2 PO 4 (1g / L), K 2 HPO 4 (1g / L), MgSO 4 ·7H 2 O(0.25g / L), CaCl 2 (10mg / L),FeSO 4 ·7H 2 O(10mg / L),MnSO 4 ·H 2 O (0.1mg / L), ZnSO 4 ·7H 2 O (1mg / L), CuSO 4 ·5H 2 O(0.2mg / L),NiCl 2 ·6H 2 O (20μg / L), biotin (0.4mg / L), MOPS (42g / L) (pH7.4)), glucose (10g / L) and tetracycline (12.5mg / L) were added to the medium, at 30 Cultivate Corynebacterium glutamicum engineering strain SY10 under the condition of ℃ and 200rmp, when OD 600 When it reaches 0.6-0.8, add IPTG, the final concentration is 1mM, reduce the shaker speed to 120rmp, and induce fo...

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Abstract

The invention discloses a corynebacterium glutamicum engineering strain for biosynthesis of rare sugar, and a building method and application thereof, and discloses a corynebacterium glutamicum recombination strain SY10. An experiment proves that rare ketose and deoxidized ketose can be synthesized by adopting a plurality of hydroxyaldehydes and glucoses as substrates by the strain in a fermentation manner, for example, D-erythrulose can be synthesized by adopting formaldehyde and glucose as substrates, L-xylulose can be synthesized by adopting glycolaldehyde and glucose as substrates, D-sorbose and D-psicose can be synthesized by adopting D-glyceraldehyde and glucose as substrates, L-fructose can be synthesized by adopting L-glyceraldehyde and glucose as substrates, and 3R, 4S, 5R, 6R-heptose and 3R, 4R, 5R, 6R-heptose can be synthesized by adopting D-erythrose and glucose as substrates. Therefore, the corynebacterium glutamicum recombination strain SY10 disclosed by the invention can be applied to the field of production of the rare ketose and the deoxidized ketose by whole-cell fermentation; the produced rare ketose and deoxidized ketose have broad application prospects in the industries such as a food, a medicine and the like.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a Corynebacterium glutamicum engineering strain prepared by genetic engineering technology and microbiological methods and its construction method, and its use in fermentation to produce rare ketose and deoxyketose in the application. Background technique [0002] Rare sugar (RareSugar) is a type of monosaccharide and its derivatives that exist in nature but are extremely low in content (defined by the International Rare Sugar Society ISRS in 2002). Although the content of rare sugar is very small in nature, it has the advantages of low calorie, high stability, harmonious sweetness, no hygroscopicity, no cariogenicity, and high tolerance, and has attracted wide attention in the fields of diet, health care, and medicine. Concern, they can be used as additives to improve the physical and chemical properties of food, and improve the physiological and health functions of food. In addit...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/21C12N15/77C12P19/02C12R1/15
Inventor 孙媛霞杨建刚朱玥明李季涛马延和
Owner 天津怡和生物科技有限责任公司
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