A strategy to improve the production performance of polylactic acid by regulating cell morphology

A technology of polylactic acid and cells, which is applied in the field of bioengineering, can solve the problems of difficult downstream separation, long time consumption, and large pollution, and achieve good application prospects, low strain growth load, and fewer system components.

Active Publication Date: 2022-01-11
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the methods of producing polylactic acid mainly include controlling the ratio of pathway enzymes, adjusting the balance of cofactors, and predicting knockout targets by the model. The content can reach up to 49%, but there are problems of low content, difficult downstream separation, long time consumption, and large pollution.

Method used

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  • A strategy to improve the production performance of polylactic acid by regulating cell morphology
  • A strategy to improve the production performance of polylactic acid by regulating cell morphology
  • A strategy to improve the production performance of polylactic acid by regulating cell morphology

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Embodiment 1

[0037] Example 1 Evaluation of Cell Morphology

[0038]The sulA gene encoding the cell division inhibitor sulA, the minC gene encoding the septum-forming protein (nucleotide sequence shown in SEQ ID NO.8), the minD gene (nucleotide sequence shown in SEQ ID NO.9), minE gene (nucleotide sequence as shown in SEQ ID NO.10), mreB gene encoding cytoskeletal protein (nucleotide sequence as shown in SEQ ID NO.11), the ATG position in front of the gene was respectively adopted in the way of fusion PCR Add B0034RBS. The above PCR products were recovered and ligated with the vector pSC101 plasmid by enzyme digestion to obtain recombinant plasmids pSC101-sulA, pSC101-minC, pSC101-minD, pSC101-minE, pSC101-mreB, respectively.

[0039] The obtained recombinant plasmids pSC101-sulA, pSC101-minC, pSC101-minD, pSC101-minE, and pSC101-mreB were respectively introduced into competent cells E. coli JM109 to obtain plasmids containing the cell division inhibitor sulA.

[0040] In LB medium, the ...

Embodiment 2

[0041] The detection of polylactic acid content in the shake flask of embodiment 2

[0042] The already constructed expression polylactic acid synthesis pathway ( figure 2 ) Carrier P of related genes J23119 -phaABC-pCT and pSC101-sulA( image 3 ) into the competent cells of the lactic acid production strain GL0002 to obtain J23119 - Polylactic acid producing strains of phaABC-pCT and pSC101-sulA plasmids.

[0043] The above-mentioned polylactic acid production strains were cultured in NBS medium, 37°C, 200rpm, and the initial OD of strain fermentation was 600 0.04, fermented for 85h. Identify the content. The result is as Figure 4 As shown, with the prolongation of bacterial culture time, the content of intracellular polylactic acid can increase to 24.2w%, the dry weight can also increase to 8.12g / L, and the glucose consumption increases gradually. At the same time, it can be seen that due to the elongation of the cells, the polylactic acid content in the cells incre...

Embodiment 3

[0045] The detection of polylactic acid content in the fermentor of embodiment 3

[0046] The fermentation performance of the DS7 strain was tested in a 7.5L fermenter filled with NBS medium. Liquid volume is 3.5L, pH is 6.7, pressure is 1mpa, temperature is constant at 37°C, 500rpm, initial inoculation OD 600 is 0.2, the ventilation rate is 1vvm, and the fermentation cycle is 84h.

[0047] Such as Figure 5 As shown, at the end of the fermentation, the accumulated content of polylactic acid reached 53.8w%, and the dry weight reached 27.2g / L.

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Abstract

The invention discloses a strategy for regulating cell morphology to improve the production performance of polylactic acid, which belongs to the technical field of bioengineering. The present invention utilizes the cell division inhibitor to inhibit cell division by means of molecular biology, so as to increase the volume and specific surface area of ​​cells and facilitate the accumulation of intracellular products. By introducing a regulatory circuit targeting cell division inhibitors, the volume of the cells is enlarged, resulting in more intracellular accumulation of polylactic acid. The content of the polylactic acid produced by the invention reaches 53.8w%, and the dry weight reaches 27.2g / L.

Description

technical field [0001] The invention relates to a strategy for regulating cell morphology to improve the production performance of polylactic acid, which belongs to the technical field of bioengineering. Background technique [0002] Polylactic acid is a good biocompatible material. Polylactic acid is a polymer obtained by polymerizing lactic acid as the main raw material. The raw material source is sufficient and can be regenerated. The production process of polylactic acid is non-polluting, and the product can be biodegraded to realize the cycle in nature, so it is an ideal green polymer material. [0003] The dynamic control system is an emerging metabolic flow control method in the field of metabolic engineering. Its main feature different from the static control is that during the fermentation process, the engineered strains will adjust according to the fermentation time, physiological state, intracellular metabolite concentration, and changes in the extracellular envir...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/21C12P7/62C12R1/19
CPCC07K14/245C12N15/52C12N9/0071C12N9/0006C12N9/1029C12N9/13C12P7/625C12Y114/14C12Y101/01036C12Y208/03001
Inventor 刘立明丁强陈修来李洋刘佳罗秋玲
Owner JIANGNAN UNIV
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