Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Engineering strain capable of producing succinic acid at low pH value and method for producing succinic acid by fermentation

A technology for producing succinic acid and genetically engineered bacteria is applied in the field of bioengineering to achieve the effect of promoting progress and development

Inactive Publication Date: 2014-12-24
NANJING TECH UNIV
View PDF3 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Escherichia coli is widely used in industry, but most Escherichia coli cannot ferment and produce succinic acid at low pH because they cannot tolerate low pH

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Engineering strain capable of producing succinic acid at low pH value and method for producing succinic acid by fermentation
  • Engineering strain capable of producing succinic acid at low pH value and method for producing succinic acid by fermentation
  • Engineering strain capable of producing succinic acid at low pH value and method for producing succinic acid by fermentation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] (1) Using LB medium, cultivate Escherichia coli AFP111 to OD at 37°C under aerobic conditions 600 = 0.5~0.6, using CaCl 2 prepared by the method lacking the lactate dehydrogenase gene ( wxya ), pyruvate formate lyase gene ( PPML ) activity and the phosphotransferase system ptsG Escherichia coli AFP111 competent strain with spontaneous mutation of gene chromosome;

[0037] The formula of LB medium was: peptone 10 g / L, yeast powder 5 g / L, NaCl 5 g / L. (2) Synthesize primers without restriction sites,

[0038] Upstream primer: 5'-ACACGAGTCCTTTGCACTTGCTTACTTT-3';

[0039] Downstream primer: 5'-CGCTCCCTTGTCTTATAACCATTCAGAC-3'.

[0040] extract E. coli JM109 (preserved in this laboratory) genome, with E. coli The JM109 genome was used as a template, and the target gene fragment was amplified by PCR. The specific reaction conditions were: 94 °C for 5 min; (94 °C for 45 s, 60 °C for 45 s, 72 °C for 4 min, 36 cycles); 72 °C for 10 min. Purified and amplified conta...

Embodiment 2

[0046] The Escherichia coli BA600 constructed in Example 1 was used as the starting strain. 1% (v / v) inoculum was transferred from the cryopreservation tube to the test tube, cultivated overnight at 37°C and 200 rpm, and then transferred to the Erlenmeyer flask with 1% (v / v) inoculum, and cultivated at 37°C and 200 rpm6 -8 h to obtain the bacterial solution in the logarithmic growth phase; inoculate the bacterial solution in the logarithmic growth phase into a 500 mL continuous culture device with 300 mL of fermentation medium at an inoculation amount of 10% (v / v), and place it in a water bath at 37 °C. Heat, pass through filter-sterilized CO 2 Maintain the anaerobic environment, and add fresh fermentation medium to the culture device at a rate of 1.5 mL / h. Regularly take samples to detect the density of bacteria in the culture device, when the density of bacteria in the reactor reaches OD 600 = 2 to 3, and there was no significant change for 48 h, indicating that the bacter...

Embodiment 3

[0051] Different carbonates with different molar concentrations were added to the fermentation medium, and CO was passed for 2 min before and after sterilization. 2 After the final pH value, the carbonates that can adjust the pH value to around 5.6 are screened out.

[0052] The formula of the fermentation medium is: citric acid 3 g L -1 , Na 2 HPO 4 12H 2 O 4 g L -1 , KH 2 PO 4 8 g L -1 , (NH 4 ) 2 HPO 4 8 g L -1 , NH 4 Cl 0.2 g L -1 , (NH 4 ) 2 SO 4 0.75 g·L -1 , MgSO 4 ·7H 2 O 1 g L -1 , CaCl 2 2H 2 O 10.0 mg·L -1 , ZnSO 4 ·7H 2 O 0.5 mg L -1 , CuCl 2 2H 2 O 0.25 mg L -1 , MnSO 4 ·H 2 O 2.5 mg L -1 , CoCl 2 ·6H 2 O 1.75 mg L -1 , H 3 BO 3 0.12 mg·L -1 , Al 2 (SO4 ) 3 1.77 mg·L -1 , Na 2 MoO 4 2H 2 O 0.5 mg L -1 , ferric citrate 16.1 mg·L -1 , 20.0 mg L -1 VB 1 , 2.0 mg L -1 Biotin, glucose 30-40 g / L.

[0053] Table 1 Comparison of pH adjusted by different carbonates obtained by screening

[0054]

[0055] The pH reg...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a genetic engineering strain capable of producing succinic acid at a low pH value. The classification name of the genetic engineering strain is Escherichia coli BA601, and the collection number is CCTCCNO: M2014210. The invention further discloses a method for producing succinic acid by using the strain. According to the strain disclosed by the invention, an Escherichia coli AFP111 which is lack of lactate dehydrogenase (LDH) genes and has activity of pyruvate formate lyase (PFL) genes and ptsG spontaneous mutation of a phosphotransferase system is taken as a starting strain for over-expression of acid-resistant genes gadBC. The method of enabling the Escherichia coli AFP111 which can not grow under low-pH (pH 5.6) conditions originally and can not produce acid by fermentation to efficiently utilize glucose to produce succinic acid through a biological and microbial acclimation means is not disclosed before, and the application can greatly promote the progress and development of the succinic acid industry.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and relates to a succinic acid-producing genetically engineered bacterium and a method for fermenting and producing succinic acid, specifically a recombinant bacterial strain producing succinic acid at a low pH value and using the bacterial strain to ferment and produce succinic acid. diacid method. Background technique [0002] Succinic acid, also known as succinic acid, is widely used in industries such as medicine, pesticides, dyes, spices, paints, food and plastics. As a C4 platform compound, it can be used to synthesize 1,4-butanediol, tetrahydrofuran, Organic chemicals such as γ-butyrolactone and biodegradable materials such as polybutylene succinate (PBS) are considered by the US Department of Energy to be among the 12 most valuable biorefinery products in the future. In addition, succinic acid is also used in the production of pharmaceuticals, antibiotics, amino acids and vitamin...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/21C12N15/70C12P7/46C12R1/19
Inventor 姜岷陈吴方吴明科马江锋刘嵘明陈可泉韦萍欧阳平凯
Owner NANJING TECH UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com