NKG mice are a type of severe immunodeficient mouse developed by Cyagen by knocking out the Il2rg gene in the NOD-Scid background strain. This strain lacks mature T, B, and NK immune cells, has reduced complement activity, and weak phagocytic activity of macrophages against human cells. Therefore, NKG mice can efficiently engraft human hematopoietic stem cells (HSC), peripheral blood mononuclear cells (PBMC), patient-derived xenografts (PDX), or adult stem cells and tissues. In the field of immunology, there are differences between humans and mice in terms of physiology and the immune system. Therefore, research conducted directly on mice does not fully reflect the situation in humans. By transplanting human peripheral blood mononuclear cells (PBMCs) or human hematopoietic stem cells (HSCs) into immunodeficient mice, the mouse’s immune system can be partially or completely replaced by the human immune system, which can simulate the function of the human immune system in vivo and provide an effective model for studying the human immune system. Transplanting PBMCs into NKG mice has the advantages of high immune reconstitution efficiency and fast speed. After 3 weeks of transplantation, the average proportion of human CD45+ cells in peripheral blood exceeds 40%. Due to individual differences in donor cells, the reconstitution efficiency of some PBMC donors in NKG mice may be higher. However, due to the mismatch between human immune cells and mouse major histocompatibility complex (MHC) molecules, graft-versus-host disease (GvHD) occurs, and transplanted human immune cells (including T cells, B cells, NK cells, etc.) attack mouse tissues, causing inflammation and tissue damage, ultimately leading to rapid death of mice. This results in a very limited experimental window period. In mice, MHC is commonly referred to as the H-2 complex. The H2-Ab1 (Histocompatibility 2, class II antigen A, beta 1) gene encodes a part of the mouse MHC class II molecules. This complex is primarily present on the surface of antigen-presenting cells such as macrophages, dendritic cells, and B cells. It plays a crucial role in presenting fragments of foreign substances, such as bacteria or viruses, to helper T cells (CD4+ T cells). This antigen presentation is a fundamental step in initiating adaptive immune responses. Studies have shown that transplanting PBMCs into MHC II-deficient mice can prevent lethal graft-versus-host disease (GvHD) caused by CD4+ T cells ^[1-2]. The absence of H2-Ab1 can affect the recognition of mouse MHC molecules by human T cells, thereby mitigating GvHD and extending the experimental window for PBMC immune reconstitution. NKG-H2-Ab1 KO mice are generated by knocking out the H2-Ab1 gene on the background of NKG mice. Compared to NKG mice, NKG-H2-Ab1 KO mice can effectively reconstitute CD8+ T cells and exhibit improved survival after PBMC transplantation. They can be used for long-term studies on PBMC immune system reconstitution.

Mmrn1 is located on chromosome 6 of mice. SgRNA and ssDNA will be designed using Nuclease Technology; Mmrn1 conditional knockout mice will be obtained by high-throughput electroporation of fertilized eggs. After sexual maturity, sperm will be collected for cryopreservation.

Dcun1d5 is located on chromosome 9 of mice. Nuclease Technology will be used to design sgRNA; Dcun1d5 knockout mice will be obtained by applying high-throughput electroporation of fertilized eggs. After sexual maturity, sperm were collected for cryopreservation.

NKG mice are a type of severe immunodeficient mouse developed by Cyagen by deleting the Il2rg gene from the NOD-Scid strain. This strain lacks mature T, B, and NK cells, has reduced complement activity, and weak macrophage phagocytosis of human cells. As a result, NKG mice can efficiently engraft human hematopoietic stem cells (HSC), peripheral blood mononuclear cells (PBMC), patient-derived xenografts (PDX), or adult stem cells and tissues. In the field of immunology research, there are differences between humans and mice in terms of physiology and immune systems, so research conducted directly on mice cannot fully reflect the human situation. By transplanting human peripheral blood mononuclear cells (PBMC) or human hematopoietic stem cells (HSC) into immunodeficient mice, the mouse’s immune system is partially or completely replaced by the human immune system, allowing for the simulation of human immune system function in vivo. PBMC transplantation into NKG mice has the advantages of high immune reconstitution efficiency and fast speed. However, due to the mismatch between human immune cells and mouse MHC molecules, graft-versus-host disease (GvHD) occurs, where transplanted human immune cells (including T cells, B cells, and NK cells) attack mouse tissues, causing inflammation and tissue damage, ultimately leading to rapid death of the mouse. The mouse MHC, termed the H-2 complex, is located on chromosome 17 and encodes a suite of molecules fundamental to antigen processing and T cell-mediated immunity. These are broadly categorized into class I (e.g., H2-K1 and H2-D1), expressed ubiquitously on nucleated cells and specialized in presenting endogenous antigens to CD8+ cytotoxic T cells, and class II (e.g., H2-Ab1), predominantly expressed by professional antigen-presenting cells (APCs) such as macrophages, B cells, and dendritic cells, responsible for presenting exogenous antigens to CD4+ helper T cells ^[1–2]. Previous studies have demonstrated that dual knockout (dKO) of mouse MHC class I and II molecules mitigates GvHD by abrogating xenogeneic antigen presentation, thereby enabling sustained human immune cell engraftment ^[3–5]. NKG-MHC-dKO(H2-K1/H2-Ab1/H2-D1) mice, generated through targeted deletion of key MHC class I (H2-K1, H2-D1) and class II (H2-Ab1) molecules, effectively circumvent the immunological mismatch that triggers graft-versus-host disease (GvHD). This strategic ablation substantially delays GvHD onset and supports extended in vivo observation of human immune system reconstitution following PBMC engraftment. The model provides a high-resolution platform for longitudinal interrogation of human immune cell functionality and cross-species immune dynamics within a mouse host.

H2-Ab1 is located on chromosome 17 of mice. Nuclease Technology will be used to design sgRNA; H2-Ab1 knockout mice will be obtained by applying high-throughput electroporation of fertilized eggs. After sexual maturity, sperm were collected for cryopreservation.

H2-Ab1 is located on chromosome 17 of mice. Nuclease Technology will be used to design sgRNA; H2-Ab1 knockout mice will be obtained by applying high-throughput electroporation of fertilized eggs. After sexual maturity, sperm were collected for cryopreservation.

H2-Ab1 is located on chromosome 17 of mice. SgRNA and ssDNA were designed using Nuclease Technology; H2-Ab1 conditional knockout mice were obtained by high-throughput electroporation of fertilized eggs. After sexual maturity, sperm were collected for cryopreservation.