Polypeptide, vaccine composed of polypeptide and application of polypeptide
A technology of vaccines and recombinant vectors, applied in the biological field, can solve side effects and other problems, achieve the effects of reducing senile plaques and Aβ levels, improving cognitive function damage, and good prospects for the treatment of AD
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Embodiment 1
[0042] The preparation of embodiment 1 polypeptide
[0043] (1) Insert the AOE1 epitope polypeptide into the C-terminus of the Aga2 protein for whole gene synthesis, wherein the gene of the Aβ1-15 fragment is used as a positive control, and the pCTCON2 empty vector is used as a negative control;
[0044] The nucleic acid sequence of the AOE1 epitope polypeptide is shown in SEQ ID NO.2, and the nucleic acid sequence shown in SEQ ID NO.2 is as follows:
[0045] SEQ ID NO.2:
[0046] CGT CCG GAT CAG GTG ATG TGG GAT AGC AAA CGT CCG
[0047] The pCTCON2 plasmid was then digested with NdeI and XhoI, and the digested vector pCTCON2 was fused with the AOE1 epitope polypeptide to obtain a recombinant vector.
Embodiment 2
[0048] The preparation of embodiment 2 polypeptide
[0049] Compared with Example 1, except that the copy number of the AOE1 epitope polypeptide is increased, and the copy number of the AOE1 epitope polypeptide is 7, other conditions are the same as Example 1.
[0050] The effect of the prepared polypeptide is similar to that of Example 1, and subsequent experiments were carried out using the polypeptide prepared in Example 1.
Embodiment 3
[0051] The preparation of embodiment 3 vaccines
[0052] (1) Preparation of yeast competent cells
[0053] Saccharomyces cerevisiae EBY100 was inoculated in 50mL YPD liquid medium at a ratio of 1:100, cultured with shaking at 30°C and 260rpm for 4-6h to the pre-logarithmic phase; centrifuged at 6000rpm at room temperature for 10min; discarded the supernatant and resuspended with 25mL of resuspension buffer, transferred Put it into a 250mL shake flask, shake it at 30°C for 10min; centrifuge at 6000rpm at room temperature for 10min, discard the supernatant; resuspend in 25ml of sterile water, centrifuge at 6000rpm at room temperature for 10min, discard the supernatant; Competent cells;
[0054] (2) Yeast cell transformation and identification
[0055] Take 1 μL of the constructed expression plasmid and add it to 100 μL of yeast competent cells, incubate on ice for 5 min, and perform electroporation. Immediately after electroporation, 1 mL of YPD medium was added, transferred ...
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