The Research Of The Biological Function And Mechanism Of Toe1 Gene In Mouse Preimplantation Embryo

Early embryonic developmental block is a common problem in somatic cell cloning,assisted reproduction and animal embryo engineering.It is very important to analyze the causes and mechanisms of developmental block to improve the success rate.To this end,we conducted transcriptome analysis on mouse cloned embryos from different donor sources,and screened the differentially expressed gene Toe1.Toe1 encodes a non-canonical deadenosine enzyme,which participates in the trimming of the tail of Terc poly(A),the RNA component of telomerase,and its biallelic mutation can lead to type 7 pontine cerebellar hypoplasia.However,whether Toe1 has an important function in early embryonic development has not been reported yet.This project studies the biological function and mechanism of Toe1 in mouse preimplantation embryo development.This paper mainly uses RNAi technology to inject Toe1 gene small interfering RNA(si RNA)into mouse fertilized eggs to obtain Toe1 knockdown embryos,and in vitro culture found that the embryos develop to the initial stage of blastocysts,which seems to be delayed.The results of cell incubation experiments show The blastocysts in the knockdown group could not break through the zona pellucida and hatch smoothly.Knockdown group and experimental group blastocyst stage blastomere count,cell proliferation experiment,and differential expression of p21(Cyclin Dependent Protein Kinase Inhibitor 1A)showed that Toe1 gene knockdown in embryos affect embryo cell division;Gene Trp53 m RNA,TUNEL apoptosis detection and DNA damage marker product ?-H2 AX fluorescence signal together indicate that embryos in the Toe1 knockdown group increase cell apoptosis at the blastocyst stage;the inner cell mass marker protein OCT4 and the trophoectoderm marker protein CDX2 The immunofluorescence results showed that the expression of lineage marker proteins in the Toe1 knockdown group were significantly reduced,and the results showed that Toe1 would affect the lineage differentiation of blastocysts.This thesis analyzes the effect of Toe1 on mouse early embryonic development from the perspective of cell division and differentiation,and concludes that Toe1 affects mouse preimplantation embryo development through cell proliferation,apoptosis,and lineage differentiation;Toe1 affects cell proliferation,cell proliferation,and cell lineage differentiation.The regulation of apoptosis is probably mediated by DNA damage.Experimental results prove that Toe1 gene is necessary for early embryonic development in mice.