动物造模,干细胞,新冠小鼠模型 z-bio 2021-11-03 416

　　In order to win the battle against the new crown pneumonia epidemic, it is urgent to prepare animal models for drug development and vaccine testing of new crown pneumonia. However, the most commonly used model animal mouse is not susceptible to SARS-CoV-2 because its viral receptor ACE2 is quite different from humans. Various methods are needed to make it susceptible, such as expressing human ACE2 in mice. (HACE2), because the expression of ACE2 has cell type specificity (REF), generally only gene knock-in methods can achieve accurate simulation of cell type specificity. Zheng Yongtang’s research team from Kunming Institute of Zoology, Chinese Academy of Sciences, through collaborative research with Biological Island Laboratory and Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences, reported a rapid and large-scale preparation of hACE2 gene knock-in mice and used to construct a new crown infection model method. This method directly uses tetraploid compensation technology to prepare gene-edited mice in batches directly from stem cells, shortening the current modeling time that requires half a year using conventional technology to 35 days. This technology is expected to play an important role in the biosafety emergency response system. Related research results were published in the National Science Review with the title "Rapid generation of ACE2 humanized inbred mouse model for COVID-19 with tetraploid complementation".

　　 Mouse embryonic stem cells have developmental pluripotency, and the use of tetraploid compensation technology can directly develop from mouse embryonic stem cells into complete mouse individuals. Compared with the conventional techniques for preparing gene-edited mice such as blastocyst chimerism and fertilized egg injection, this technical route has the advantage that it can bypass the breeding steps and quickly produce a large number of homozygous precision gene-edited mice, but it is also limited by four. Ploidy compensation technology is difficult to operate, and embryonic stem cell culture technology. At present, the success rate of producing tetraploid mice from inbred background or multiple passages of embryonic stem cells is extremely low. Chen Jiekai's research group in the Biological Island Laboratory has many years of experience in basic research and operation of embryonic stem cells, which can maintain the developmental stemness of inbred embryonic stem cells for a long time, and use CRISPR-Cas9 technology to edit mouse embryonic stem cells by human ACE2 insertion; Wu Guangming's research group at Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences has long been engaged in embryonic development research. In this report, its optimized mouse tetraploid compensation technology was used to quickly prepare effective C57BL/6 and BALB/c inbred strains. Human-derived ACE2 is inserted into the mouse model at a specific point, and the efficiency can reach 30%; Zheng Yongtang's group from Kunming Institute of Zoology has long been engaged in virus immunology and infection animal model research. The teams worked together to knock the human ACE2 gene into the mouse Ace2 site in situ to prepare a humanized ACE2 mouse model with the correct expression pattern, high efficiency of new coronary pneumonia virus infection, and detection of the virus in the stool. It is used to evaluate the effect of neutralizing antibodies. This research not only produced an effective humanized ACE2 mouse model for the study of new coronary pneumonia disease, but also developed a set of methods that can quickly prepare a large number of accurate gene-edited mouse models, which can quickly respond to emergencies and achieve complex Preparation of multi-gene simultaneous editing mice. Since preparing the first batch of mice in March, the team has provided more than 340 mice for drug screening and antibody neutralization experiments in many domestic units.

　　 The research was funded by the key special projects of the National Key Research and Development Program, and the emergency research projects of Guangdong Province and the Biological Island Laboratory.