Goose astrovirus spike protein liposome vaccine as well as preparation method and application thereof
A technique for preparing astroviruses and vaccines, applied in liposome delivery, virus/bacteriophage, biochemical equipment and methods, etc., can solve the problems of vaccines preventing goose astrovirus infection, etc., and achieve the effect of prolonging the action time
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Embodiment 1
[0030] Example 1 Prokaryotic expression and purification of antigenic protein VP27
[0031] The prokaryotic expression and purification process of antigenic protein VP27 is as follows: figure 1 shown. The Goose Astrovirus DY-19 strain isolated and preserved from the Pathogenic Microbiology and Immunology Research Laboratory of Anhui Agricultural University was selected, inoculated on LMH cells and cultured to the F3 generation, the cell pellet was collected, and the RNA of the Goose Astrovirus was extracted by the Trizol method , using Beyotime's M-MLV reverse transcription kit to reverse-transcribe the extracted RNA into cDNA, and store it at -20°C for later use.
[0032] The goose astrovirus DY-19 strain was designed according to a goose astrovirus (accession number: MG882765.1) on GenBank and a pair of specific primers were used to amplify the VP27 gene, as follows:
[0033] GAstV-F: 5'-CGGGATCCCAGGTTACTCCCTCGCTTGTG-3',
[0034] GAstV-R: 5'-CCCAAGCTTAGAGGTCTTGAGCGAGACTGC...
Embodiment 2
[0036] Embodiment 2 prepares blank liposome
[0037] Accurately weigh 1.13g of soybean lecithin, 0.57g of cholesterol, and 0.032g of vitamin E, dissolve them in 20mL of chloroform containing 1% (V / V) ethanol, and accelerate the dissolution of the membrane material in a horizontal shaker at 37°C. Add the dissolved mixed solution into a 250mL round bottom flask, and connect the flask to a rotary evaporator. Turn on the decompression pump, and remove the organic solvent by rotary evaporation at 42°C and 180 r / min under reduced pressure, forming a layer of homogeneous film on the bottom of the bottle, and continue the rotary evaporation under reduced pressure for 30 minutes to remove the remaining solvent to obtain a membrane-like blank liposome.
Embodiment 3
[0038] Example 3 Hydration
[0039] Add 20 mL of phosphate buffer with a pH value of 7.4 to the above-mentioned round bottom flask, and add small glass beads at the same time to accelerate the hydration process and reduce the particle size of the liposomes. Place on a horizontal shaker and hydrate at 200 r / min at 37°C for 1 hour to completely dissolve the liposomes from the wall and form a white suspension. Separate the resulting suspension into a beaker.
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