ACE2 cell humanized mouse model as well as construction method and application thereof
A mouse model and construction method technology, applied in the field of animal genetic engineering, can solve the problems of unfavorable reconstruction of the immune system, hindering the study of viruses and immune cells, long modeling cycle, etc., to reduce background pollution, avoid xenograft rejection, shorten the The effect of the experimental cycle
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Embodiment 1
[0054] The present invention provides a mouse model of ACE2 cell humanization, and its construction method is as follows:
[0055] (1) Construction of BGC823-ACE2-GFP stable cell line:
[0056] ①Use lentiviral vectors to construct plasmids overexpressing ACE2 receptor and green fluorescent protein (GFP), and then use 293T cells to prepare lentiviral supernatant suspension;
[0057] ② After centrifuging and filtering the lentiviral supernatant, use Jurkat cells to detect virus titer; among them, titer (TU / mL) = cell number × fluorescence percentage × 10 3 / Volume of virus stock solution (μL);
[0058] ③ Taking the multiplicity of infection (MOI) as 10, every 1×10 6 Add 10 to each BGC823 cell 7 TU lentivirus for infection;
[0059] ④Change the medium after 6 hours of infection, and continue to culture; 48 hours later, take the infected cells and test the infection efficiency by flow cytometry; the infection efficiency test results are as follows: figure 1 As shown, among th...
Embodiment 2
[0070] This embodiment is beneficial to step (1) in Example 1 to construct cells overexpressing ACE2 receptors with Jurkat, NALM6, Huh7, A549 and H1299 as parent cells;
[0071] The infection efficiency and the infection efficiency after sorting are shown in Table 1.
[0072] Table 1
[0073] cell name infection efficiency Infection efficiency after sorting ACE2-A549 cells 90％ --- ACE2-Huh7 cells 90％ --- ACE2-H1299 cells 44％ 90％ ACE2-Jurkat cells 50％ 90％ ACE2-Nalm6 cells 17％ 90％
[0074] It can be seen from the above table that when A549 and Huh7 are used as mother cells, the infection efficiency can reach 90%. Although the infection efficiency of the other three types of cells does not reach 90%, the infection efficiency can reach 90% after sorting.
Embodiment 3
[0076] In this example, the distribution of ACE2-positive cells in the lungs of mice was observed by pathological detection methods.
[0077] After the mice were killed, the lungs of the mice were taken and photographed under a microscope to observe the GFP signal; then a part of the lungs was used to prepare a single cell suspension, and the proportion of BGC823 cells in the lungs of the mice was detected by flow cytometry, and the other A part was used to prepare paraffin sections for HE staining and immunofluorescence detection.
[0078] The results of flow cytometry were as Figure 5 As shown, it is shown that BGC823-ACE2-GFP cells account for 27.3% of the total number of cells in the lungs of mice; wherein the negative control group is a blank mouse (no injection of cells), wherein the GFP content is 2.81%;
[0079] HE staining results as Figure 6 As shown, it shows that after the injection of BGC823-ACE2-GFP cells, tumor lesions were found in the lungs of the mice, an...
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