Recombinant expression vector, host and construction method thereof, and animal model construction method

Abstract

The invention relates to the field of biotechnology, in particular to a recombinant expression vector, a host and a construction method thereof, and an animal model construction method. The recombinant expression vector consists of nucleotide for encoding soluble humanized HLA-A2 proteins and a prokaryotic expression vector. By virtue of the special prokaryotic expression vector, 'sufficient' soluble humanized HLA-A2 proteins can be synthesized; by taking the purified HLA-A2 proteins as an immunogen, the 'accuracy' of an experiment is guaranteed; the HLA-A2 proteins at different concentrations can promote a recipient mouse to generate preformed HLA-A2 antibodies at different titers, so that the generating strength of the preformed antibodies becomes 'controllable'; and the change of 'lymphocytes in marrow and spleen' of a preformed antibody animal model can be successfully observed.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a recombinant expression vector, a host and a construction method thereof, and a construction method of an animal model. Background technique [0002] A kidney transplant is the best treatment for end-stage renal disease. With the widespread use of immunosuppressive drugs, T cell-mediated rejection has been significantly reduced, but acute and chronic antibody-mediated rejection plays an increasingly important role in kidney allograft loss. Pre-existing anti-donor HLA (human leukocyte antigen) antibodies are closely related to graft failure and antibody-mediated rejection. The success rate of kidney transplantation in patients with HLA hypersensitivity is low, so effective desensitization therapy is the key to successful kidney transplantation. Animal experiments are one of the main means of transplantation immune research, so the successful synthesis of "sufficient" soluble humaniz...

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Problems solved by technology

[0006] Disadvantages of the prior art: 1. The eukaryotic expression system has low protein expression, high cost, long time-consuming and difficult purification, which is not enough to construct a large number of pre-existing specific antibody animal models to explore antibody clearance schemes; 2. Mix MICA protein with an equal volume After Freund's complete adjuvant was mixed and injected through the tail vein, the specific anti-MICA antibody in the recipient mice was suspiciously positive, and the animal model of pre-existing anti-MICA antibody was not successfully established; 3. Insect Sf-9, which can express MICA protein After the cell suspension was injected through the tail vein, although the specific MICA antibody in the recipient mice was positive, in addition to expressing the MICA protein in this method, the Sf-9 cells can also produce a large amount of self-protein, so in the process of sensitizing the recipient mice, Inevitably, cross-reactions will occur, thereby affecting the accuracy of the experiment; 4. The current method of constructing a kidney transplant animal model with pre-existing anti-specific antibodies cannot control the titer of the pre-existing specific antibodies; nor has it been observed that the sensitization process interacts with antibodies produce related changes in lymphocytes

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