Engineering escherichia coli capable of efficiently producing hyaluronic acid and preparation method thereof

A technology of hyaluronic acid and Escherichia coli, applied in the field of genetic engineering of microorganisms, can solve the problems of low safety and shortage of hyaluronic acid resources, etc.

Active Publication Date: 2012-09-05
MICROBIAL FERMENTATION ENG RES CENT CO LTD OF YUNNAN PROVINCE
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0010] The purpose of the present invention is to provide a kind of high-efficiency hyaluronic acid-producing engineering Escherichia coli and its preparation method, to overcome the shortage of resources and safety problems in the production of hyaluronic acid in the prior art and the low hyaluronic acid synthesis in engineering Escherichia coli question

Method used

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  • Engineering escherichia coli capable of efficiently producing hyaluronic acid and preparation method thereof
  • Engineering escherichia coli capable of efficiently producing hyaluronic acid and preparation method thereof
  • Engineering escherichia coli capable of efficiently producing hyaluronic acid and preparation method thereof

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Embodiment

[0064] from Pasteurella multocida ( Pasteurella multocida subsp. Multocida ) in the hyaluronan synthase gene pmHas clone and containing the hyaluronan synthase gene pmHas Construction and transformation of the expression vector pQEpmHas Escherichia coli JM109 strain. Specific steps are as follows:

[0065] (1) Construct the expression vector pQEpmHas containing the hyaluronic acid synthase gene, and clone it from Pasteurella multocida ( Pasteurella multocida subsp. Multocida ) in the hyaluronan synthase gene ( P Asteurella m ultocida h Yaluronic a cid synthase PmHas) , the primers used for its PCR amplification clone are designed as follows:

[0066] Upstream primer pmHas P1 is 5'-TA GGATCC ATG AACACATTATCACAAGCAAT-3', (SEQ ID No:3 in the sequence listing) contains Bam HI site GGATCC and initiation codon ATG;

[0067] The downstream primer is pmHas P2 is 5'-TA GAGCTC TTA TAGAGTTATACTATTAATAAT-3' (SEQ ID No:4 in the sequence listing), cont...

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Abstract

The invention provides an engineering escherichia coli capable of efficiently producing hyaluronic acid (HA) and a preparation method thereof. The engineering escherichia coli is obtained by transferring escherichia coli through constructing recombinant expression vector commonly expressed in escherichia coli by gene Has containing HA synthetase and gene containing uridine diphosphate glucose phosphate dehydrogenate. The engineering escherichia coli capable of efficiently producing HA, provided by the invention, is pHK / JM109 with CGMCC No.3926. The engineering escherichia coli provided by the invention can produce 29-2.5g / L of HA. Compared with the yield in the prior art, the highest yield of HA obtained from gene spHas synthesized by Gram positive strep HA which is expressed in escherichia coli is enhanced by about more than 10 times.

Description

technical field [0001] The invention relates to the technical field of genetic engineering of microorganisms, in particular to a high-efficiency hyaluronic acid-producing engineering Escherichia coli constructed in the form of a plasmid or in a genome integration manner and a preparation method thereof. Background technique [0002] Escherichia coli K-12 strain is widely used in the field of metabolic engineering to produce a variety of high value-added products due to its clear genetic background, numerous efficient genetic transformation systems and the ease of genome manipulation. No natural strain of E. coli has been found to have the ability to produce hyaluronic acid. [0003] Hyaluronic acid (Hyaluronic acid, HA) is a high-viscosity amino polysaccharide that was first isolated from bull eyes. Hyaluronic acid widely exists in human and animal tissues, and its molecular weight is 5×10 4 to 8×10 6 Between Daltons, it is glucuronic acid linked by β-1,4-glycosidic bonds...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/21C12N15/70C12R1/19
Inventor 毛自朝杨发祥尚海丽程倩
Owner MICROBIAL FERMENTATION ENG RES CENT CO LTD OF YUNNAN PROVINCE
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