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Application of plants as hosts in expressing canakinumab

An expression vector and plant technology, applied in the application field of plants as hosts in the expression of kana antibody, can solve the problems of low production capacity of animal cells, complicated operation, high price, etc., and achieve the reduction of biosafety problems, simple purification, and production convenient effect

Pending Publication Date: 2018-02-23
SAGACITY FAITHFUL CONVERGENCE HEALTH TECH LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, animal cell culture requires expensive culture medium, strict factory conditions, complex operations, a time period of at least two weeks, and low production capacity of animal cells, resulting in extremely high costs
Sometimes the virus carried by animal cells can infect humans, resulting in low safety

Method used

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  • Application of plants as hosts in expressing canakinumab
  • Application of plants as hosts in expressing canakinumab
  • Application of plants as hosts in expressing canakinumab

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] The construction of embodiment 1 plant transient expression vector

[0051] In order to provide high-efficiency expression of foreign aid proteins in plants, the protein sequence reverse translation software (https: / / www.idtdna. com / CodonOpt) to optimize its codons to plant-preferred codons, which were synthesized by Kingsray Company (Nanjing, China). An Xbal restriction site was added to the 5' end of the optimized Kanal heavy chain sequence, and a SacI site was added to the 3' end. A Xbal restriction site was added to the 5' end of the kana light chain sequence, and a Sacl site was added to the 3' end. and cloned into the pUC57 vector by GenScript ( figure 1 ), generate pUC57-Cana-H, pUC57-Cana-L cloning vectors respectively. Gene fragments were isolated from cloning vectors by Xbal / Sacl, and cloned into binary plant vector, pCam35S, to generate plant expression vectors p35S-Cana-H, p35S-Cana-L, respectively. The two plant expression vectors were respectively tran...

Embodiment 2

[0053] Example 2 Agrobacterium-mediated vacuum infiltration

[0054] Mix the prepared Agrobacteria containing p35S-Cana-H and p35S-Cana-L in equal amounts until the O.D.600 is 0.5. The culture suspension was placed in a 2L beaker and placed in a desiccator. The lettuce kept in this laboratory was turned upside down (core up) and gently swirled in the bacterial suspension, and the desiccator was sealed. The vacuum pump (Welch Vacuum, Niles, IL, USA) was turned on to evacuate and the permeate was seen in the leaf tissue. Hold the pressure for 30-60 seconds. The system is quickly opened to release the pressure and allow permeate to seep into the spaces within the tissue. This process was repeated 2 to 3 times until the penetration of the permeate into the lettuce tissue was clearly visible. The lettuce tissue was then gently removed from the permeate and rinsed three times consecutively with distilled water before being transferred to a container covered with plastic film. T...

Embodiment 3

[0054] Mix the prepared Agrobacteria containing p35S-Cana-H and p35S-Cana-L in equal amounts until the O.D.600 is 0.5. The culture suspension was placed in a 2L beaker and placed in a desiccator. The lettuce kept in this laboratory was turned upside down (core up) and gently swirled in the bacterial suspension, and the desiccator was sealed. The vacuum pump (Welch Vacuum, Niles, IL, USA) was turned on to evacuate and the permeate was seen in the leaf tissue. Hold the pressure for 30-60 seconds. The system is quickly opened to release the pressure and allow permeate to seep into the spaces within the tissue. This process was repeated 2 to 3 times until the penetration of the permeate into the lettuce tissue was clearly visible. The lettuce tissue was then gently removed from the permeate and rinsed three times consecutively with distilled water before being transferred to a container covered with plastic film. The treated samples were kept in the dark for 4 days. Example 3...

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Abstract

The invention relates to the technical field of biology, in particular to application of plants as hosts in expressing canakinumab. Plants such as lettuce are adopted as an effective expression platform of recombinant protein production, and a simple and effective agrobacterium tumefaciens mediated transformation vacuum infiltration method is utilized to express canakinumab, ilaris, ACZ885, anti-human IL-1beta-monocolonal antibody. The expression system can be collected after it is determined that botanical foreign protein is infected with agrobacterium for 4 d. An SDS-PAGE method is utilizedto determine that the recombinant canakinumab is successfully expressed. As is proved by polymorphonuclear leukocyte inhibition experiments, the canakinumab produced by the lettuce has biological activities of inhibiting interleukin-1beta and inhibiting neutrophile granulocyte.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to the application of plants as hosts in the expression of kana antibodies. Background technique [0002] Cancer is the leading cause of death around the world and its incidence is rising due to the growing and aging population and the prevalence of other factors such as pollution of air, food, etc. Among the treatment methods, surgery, chemotherapy and radiation irradiation are still the main means of treating various tumor types and stages at this stage. However, chemotherapy approaches have limited success due to lack of selectivity for tumor cells and lead to systemic toxicity and drug resistance. Radiation therapy also doesn't kill all the cancer cells and makes the patient more debilitated. Because cancer cells can spread, surgery can remove the diseased part, but it can't stop the spread of cancer cells. More advanced therapies are based on the molecular characteristics of tum...

Claims

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Application Information

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IPC IPC(8): C12N15/82C07K16/24
CPCC12N15/8258C07K16/244C12N2800/22C07K2317/14
Inventor 王跃驹马洁焦顺昌
Owner SAGACITY FAITHFUL CONVERGENCE HEALTH TECH LTD
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