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Retinoic acid inducible gene Ⅰ(RIG-Ⅰ)is a critical RNA virus sensor that serves to initiate antiviral innate immunity,was identified as a pattern-recognition receptor(PRR)that mediates antiviral immunity by inducing type Ⅰ interferon production.To further study the biological function of RIG-Ⅰ,we generated RIG-Ⅰ-/-mouse models by CRISPR/Cas9 system and functional analysis,taking a different strategy to the previously reported strategy.We generated RIG-Ⅰ gene knockout mice strains by embryo co-microinjection of Cas9 mRNA and sgRNA mRNA targeting the first exon of RIG-Ⅰ gene.T7END1 enzyme digestion and sequencing results shown that succssfully generated-41bp deficiency RIG-Ⅰ+/-mice,after breeding two generations with WT female mice,then breeding RIG-Ⅰ+/-male with RIG-Ⅰ+/-female mice,to obtain RIG-Ⅰ-/-homozygous in F3 generation.We successfully achieved RIG-Ⅰ-/-mice are viable and fertile.Compared with WT mice,RT-PCR results showed that RIG-Ⅰ gene transcription is negative in the kidney,spleen,liver tissue and western-bolt results showed that RIG-Ⅰ protein is not expression in the spleen of RIG-Ⅰ-/-mice;Analysis of 3-12 weeks bodyweight suggested that RIG-Ⅰ deficiency had no significant effect on growth of mice;Automated blood analyzer analysis of peripheral blood in mice results showed that lymphocytes ratio decreased,granulocyte ratio increased;Hematoxylin-eosin staining analysis of mice spleen tissue results showed that white pulp lymphocytes mild decreased in RIG-Ⅰ-/-mice;Flow cytometry analysis RIG-Ⅰ-/-mice peripheral blood results show that the number of DC cells and B cells were decreased.The expression of MAVS,IRF3,IFN-β,TNF-a,IL-6 mRNA in spleen of RIG-Ⅰ-/-mice were decreased after Poly I:C stimulated,proved that RIG-Ⅰgene knockout affect RIG-Ⅰ signaling pathways induced type Ⅰ interferon and inflammation factor.The number of CD4+T cells is significant decreased,and CD8+T cells is increased,proved RIG-Ⅰ gene knockout influence T lymphocytes activation after Poly I:C stimulated.All these findings suggested that RIG-Ⅰ-/-mice were immunodeficiency mice and provided good animal models for the study of human diseases and animal infectious diseases prevention or treatment strategies.