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Escherichia coli engineered strain for producing allitol by virtue of biotransformation as well as construction method and application thereof

An engineering strain, alliitol technology, applied in the biological field, can solve problems such as poor bacterial substrate specificity, and achieve the effect of wide application prospects

Active Publication Date: 2014-07-30
TIANJIN INST OF IND BIOTECH CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] This patent discloses the method of using the oxidation and reduction of Enterobacter agglomerans 221e to produce a variety of rare sugars and sugar alcohols respectively, but the substrate specificity of the bacteria Poor, if converted with a mixture of D-fructose and D-psicose, other by-products may be formed

Method used

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  • Escherichia coli engineered strain for producing allitol by virtue of biotransformation as well as construction method and application thereof
  • Escherichia coli engineered strain for producing allitol by virtue of biotransformation as well as construction method and application thereof
  • Escherichia coli engineered strain for producing allitol by virtue of biotransformation as well as construction method and application thereof

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Embodiment 1

[0027] Embodiment 1, construct Escherichia coli ( Escherichia coli ) engineering strain strain II

[0028] The present invention specifically provides a strain of Escherichia coli ( Escherichia coli ) Engineering strain II and its construction method, its technical route is as follows figure 1 As shown, including the following three aspects: (1) will be derived from rumen bacteria ( Ruminococcus sp.) D-psicose 3-epimerase gene ( dpe ) was connected to the vector pET21 to construct the expression vector pET- dpe , the gene expression product can convert D-fructose into D-psicose; (2) will be derived from Klebsiella oxytoca ( Klebsiella oxytoca ) ribitol dehydrogenase gene of CGMCC7662 ( rdh ) and from Candida ( Candida methylica ) formate dehydrogenase gene of X81129 ( fdh ) linked to the vector pCDFDuet TM -1, construct the double enzyme expression vector pCDF- rdh-fdh , rdh The expression product of the gene can convert D-psicose into allicitol, fdh...

Embodiment 2

[0041] Embodiment 2, escherichia coli ( Escherichia coli ) Application of engineering strain strain II in transforming and producing allicitol

[0042] (1) Escherichia coli ( Escherichia coli ) The cultivation of engineering strain strain II

[0043] Escherichia coli ( Escherichia coli ) The engineering strain strain II CGMCC No.9024 was inoculated in LB medium containing 100 μg / ml ampicillin, 100 μg / ml streptomycin and 10 μg / ml chloramphenicol, and the strain was fermented at 37°C, pH 7.0, and 200 rpm cultured to OD 600 The value is 0.8-1.0, add IPTG to the final concentration of 1.0mmol / L, 20°C, 100rpm, induce overnight.

[0044] (2) Escherichia coli ( Escherichia coli ) Preparation of engineering strain strain II quiescent cells

[0045] The induced Escherichia coli were collected by centrifugation ( Escherichia coli ) engineering strain II CGMCC No.9024 bacteria, wash the bacteria with pH 7.0 Tris-HCl buffer solution (recipe: 50mM Tris aqueous solution, adjus...

Embodiment 3

[0048] Embodiment 3, escherichia coli ( Escherichia coli ) Application of engineering strain strain II in transforming and producing allicitol

[0049] The method is the same as that of Example 2, except that the reaction is carried out at 42°C for 18 hours at different pHs (4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10.0). The results showed that Escherichia coli ( Escherichia coli ) engineering strain strain II CGMCC No.9024 transforms the optimal reaction pH value of producing allicitol to 7.0, and when the pH is greater than 8.0, the transformation efficiency of the transformation reaction decreases rapidly ( image 3 ).

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Abstract

The invention provides an escherichia coli engineered strain for producing allitol by virtue of biotransformation as well as a construction method and application of the strain, and discloses an escherichia coli recombinant strain II. Experiment shows that D-fructose can be transformed into allitol by virtue of resting cell reaction of the strain, and the rare allitol is wide in application prospect in food and medicine industries.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to an engineering strain of Escherichia coli prepared by using genetic engineering technology and microbiological methods and its construction method, and its application in transforming and producing allicitol. Background technique [0002] Rare sugar (Rare Sugar) is a kind of monosaccharide and its derivatives that exist in nature but are very low in content (defined by the International Rare Sugar Society ISRS in 2002). Its taste is similar to sucrose, but it has low calories, high stability, sweet It has the advantages of harmonious taste, no hygroscopicity, no cariogenicity, and high tolerance, which plays an important role in improving the diet of special populations. In addition, a large number of research results also show that rare sugars play an important physiological role in many aspects such as anti-cancer, scavenging free radicals, neuroprotection, etc., and can also under...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N15/70C12P7/18A61K31/047A61P3/10A23L1/29C12R1/19A23L33/00
Inventor 朱玥明李泓漪曾艳门燕孙媛霞
Owner TIANJIN INST OF IND BIOTECH CHINESE ACADEMY OF SCI
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