Analysis Of Lymphocyte Development And Function In Fosb Gene Knockout Mice

Research purpose:Our previous RNAseq data find that the Fosb gene is highly expressed in germinal center B cells compared to the na(?)ve-B cells,but the function role is still unclear.In order to further explore the specific mechanism of this gene on lymphocytes,this project plans to study the Fosb effect on the development of T and B lymphocytes and the differentiation of HSC(Hematopoietic stem cells),and on the formation of the germinal center B cells,as well as the production of antibodies,in response to immune stimulation by the Fosb gene knockout in mice.Research method:Flow cytometry(FCM),a biological technique uses to count and sort tiny particles suspended in fluids,which obtains single cells or other biological particles in suspension to detecting labeled fluorescent signals for achieving high-speed,one-by-one quantitative analysis and sorting of cells.used to After constructing the Fosb knockout mouse modelby the CRISPR/Cas9 gene editing technology,I used flow cytometry to examine and analyze the changes of T and B lymphocytes and their subgroups in varous immune organs and ascites,as well as the differentiation of hematopoietic stem cells in Fosb^+/+and Fosb^-/-mice.By injecting NP₂₀-CGG and sheep red blood cells into Fosb^+/+and Fosb^-/-mice for immune stimulation,I further examined the formation of germinal center and the production of antibodies.Research results:Fosb gene knockout mice have undergone a frameshift mutation after gene editing and can be inherited stably,and the Fosb gene has been completely knocked out as identified at the DNA,mRNA,and protein levels;Fosb^-/-is analyzed by flow cytometry and compared with Fosb^+/+mice,there is no significant difference in the differentiation of T,B lymphocytes and their subgroup cells and hematopoietic stem cells in bone marrow.The antigen NP₂₀-CGG was injected intraperitoneal to immunostimulate B cells for forming germinal centers Fosb^-/-and Fosb^+/+.However,the antigen-specific antibodies showed no significant difference after statistical analyses.Conclusion:It was verified that we successfully geneated the Fosb gene knockout mouse,as demonstrated at the DNA,mRNA,and protein levels.Comparing with wild-type,there was no significant difference in the differentiation of T,B lymphocytes and bone marrow hematopoietic stem cells in Fosb knockout mice,and neither the formation of B cell germinal centers nor the production of antigen-specific antibodies were significantly affected by knockouting the Fosb gene.