Construction And Identification Of Inducible Rbm24 Knockout Mouse

RNA-binding proteins are key players in post-transcriptional regulation by regulating alternative splicing and stability of mRNA.Rbm24(RNA-binding protein24)is a member of the RNA Binding protein family,which is conserved in di fferent species and highly expressed in heart and skeletal muscle.Previous studies demonstrated the functional role of Rbm24 in the differentiation and development of heart and skeletal muscle.Recently,it has been found that Rbm24 may also have important functions in other tissues,organs and diseases.The function of Rbm24 can be further studied by constructing Rbm24 knockout mice.However,Rbm24 knockout mice are embryorucally-lethal.In order to investigate the role of Rbm24 in tissues,organs and diseases of adult mice and achieve Rbm24 time-specific knockout,the Cre-loxP system is used to construct tamoxifen-induced systemic Rbm24 knockout mice.The inducible Cre-loxP system can be used to knock out a gene in a specific period or in a specific tissue,and it is a powerful tool for studying the function of the gene in a specific tissue and period.Rbm24loxp/loxp mice were generated by homologous recombination,and mated with R26CreER mice,which expresses Cre recombinase after induced by Tamoxifen,to obtain F1 double-hybrid mice.self-inbred of F1 generation mice could obtain the inducible Rbm24 systemic knockout mice.Mice with cre gene and loxP/loxP loci were selected by genotyping.And mice were intraperitoneally injected with tamoxifen to induce Rbm24 knockout in tissues and organs.To examine the feasibility and efficiency of the Cre-loxP knockout system,the expression of Rbm24 was detected by western blotting at different time points in different tissues.The cardiac phenotype of the inducible knockout mice was analyzed,and it was functionally verified that the Rbm24 inducible knockout mice were successfully constructed.This recombination was confirmed in the DNA of the mutant mice by genotyping.Rbm24 in mouse heart and skeletal muscle has been completely knocked out on the 3 days after injection,which confirmed the system can used to delete the target gene,as well as worded high-efficiency.Changes in the alternative splicing forms of the Rbm24 downstream genes were detected,we evaluated heart performance by cardiac histopathology and echocardiography on control mouse and iKO mouse respectively.Gross morphology and cardiac histopathology of control and iKO hearts of mice showing enlarged left ventricular chamber size and wall thinning.We also observed increased dilation of the ventricle chambers and the increased fibrosis.Echocardiography was performed to analyze cardiac function,the results showed decrease in fractional shortening(FS)and ejection fraction(EF),suggesting deteriorated cardiac function in Rbm24 iKO mice.According to the above results,we identified that the inducible knockout mice had been successfully constructed.The inducible knockout mice could be used as a significant platform to research the potential function of Rbm24.