A kind of GFP-2 small peptide and its preparation method and application

A GFP-2, culture medium technology, applied in the preparation method of peptides, biochemical equipment and methods, chemical instruments and methods, etc., can solve the problems of inability to achieve immune body efficacy, low immunogenicity, etc., and improve immune efficacy. , Improve antibody titer, strengthen the effect of specific immune response

Active Publication Date: 2021-08-17
杭州洪晟生物技术股份有限公司
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0004] The technical problem to be solved by the present invention is that the immunogenicity is often low, and the effect of the immune body cannot be achieved. To overcome the deficiency of LB medium, brown sugar medium is used to expand and cultivate GFP-2 bacteria, and a preparation of small peptide is provided. The method, the medium can increase the expression of small peptides, which is more conducive to industrial production, and the small peptides are successfully expressed and secreted into the medium, which is beneficial to the preparation and purification of small peptides, and is conducive to maintaining the activity of small peptides

Method used

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  • A kind of GFP-2 small peptide and its preparation method and application

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preparation example Construction

[0025] First of all, the present invention provides a preparation method of GFP-2 small peptide, the preparation method comprising the following steps:

[0026] (1) Prepare brown sugar medium and sterilize it;

[0027] (2) Inoculate GFP-2 bacteria, glycerol bacteria and LB medium, and culture on a shaking table;

[0028] (3) Transfer to brown sugar culture medium, carry out shaking table culture, then carry out centrifugation, collect supernatant, and discard bacteria;

[0029] (4) Salt out to precipitate proteins and small peptides, centrifuge again after salting out, remove supernatant, and collect crude protein;

[0030] (5) Resuspend the crude protein with tris-HCl, pass the resuspended crude protein through the membrane, and dialyze with tris-HCl;

[0031] (6) Perform ultrafiltration with an ultrafiltration tube to collect small peptides with a molecular weight of less than 10KD, cool the obtained small peptides, and freeze-dry to obtain GFP-2 small peptides.

[0032] ...

Embodiment 1

[0045] Preparation of GFP-2 Small Peptide

[0046] (1) Prepare 1 L of brown sugar medium and sterilize at 110°C for 15 minutes;

[0047] (2) Inoculate GFP-2 bacteria, 5mL LB medium per tube, and inoculate glycerol bacteria and medium at a ratio of 1:100. A total of 2 tubes of GFP-2 bacterial solution were connected and cultured on a shaking table for 12 hours;

[0048] (3) Transfer to 1L brown sugar medium, culture on a shaker for 12 hours, centrifuge at 4000 rpm for 1 hour, collect the supernatant, and discard the bacteria;

[0049] (4) Prepare saturated ammonium sulfate. Salt out the supernatant and saturated ammonium sulfate at a ratio of 4:1 to precipitate proteins and small peptides. After salting out, centrifuge again at 12,000 rpm for 20 minutes, remove the supernatant, and collect crude protein.

[0050] (5) Resuspend the crude protein with tris-HCL, pass the resuspended crude protein through a 0.45mm filter membrane, dialyze with tris-HCL for 5 hours, and change the...

Embodiment 2

[0053] Study of GFP-2 Small Peptide on Improving Antibody Titer

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Abstract

The present invention relates to a GFP-2 small peptide and its preparation method and application. The small peptide is prepared by the following methods: (1) preparing a brown sugar culture medium and sterilizing it; (2) inoculating with GFP-2 bacteria, glycerol bacteria and Inoculate with LB medium and culture on a shaker; (3) Transfer to brown sugar medium, culture on a shaker, then centrifuge, collect supernatant, and discard bacteria; (4) Salt out to precipitate proteins and small peptides , centrifuge again after salting out, remove the supernatant, and collect the crude protein; (5) Resuspend the crude protein with tris‑HCl, pass the resuspended crude protein through the membrane, and dialyze with tris‑HCl; The filter tube is subjected to ultrafiltration to collect small peptides with a molecular weight of less than 10KD, and the obtained small peptides are cooled and freeze-dried to obtain GFP‑2 small peptides. The prepared small peptide can be used as an adjuvant, which can greatly improve the immune efficacy.

Description

technical field [0001] The invention relates to the technical field of biotechnology and pharmaceutical engineering, in particular to a method for preparing a small peptide adjuvant derived from marine bacillus GFP-2, and the application of the small peptide in improving the immune efficacy of vaccines. Background technique [0002] Vaccination is the most cost-effective way to prevent and treat infectious diseases, and adequate immunogenicity and safety of vaccines are the most important concerns in this field. While traditional vaccines such as live, attenuated organisms and killed or inactivated organisms are generally more immunogenic, there is a limited potential risk of reversion to virulence. Additionally, these vaccines are unstable and have low safety profiles. Compared with traditional vaccines, new vaccines such as DNA vaccines, recombinant vaccines and subunit vaccines are relatively safe. However, these vaccines also have certain defects. One of the biggest de...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P21/02C07K14/32C07K1/34A61K39/39A61P37/04C12R1/07
CPCA61K39/39A61K2039/55516A61P37/04C07K14/32C12P21/02
Inventor 吕正兵舒建洪王丹
Owner 杭州洪晟生物技术股份有限公司
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