Study On Stat1b Gene Mutation On The Transcriptome Regulation Of Zebrafish Embryo In Early Development

Signal transducer and activator of transcription 1?STAT1?are associated with signal transduction of various cytokines and growth factors,and regulates many cellular functions,such as the regulation of the immune system,the inhibition of tumor cells'growth,the inhibition of cells'differentiation and the promotetion of cell apoptosis.In the previous studies,it was found that STAT1 was highly expressed in low bone mineral density?BMD?human group,and osteoclasts?OC?increased in stat1b^-/-zebrafish skeleton.The zebrafish was used for research model because of highly homologous in both genetics and physiology between zebrafish and human,strong genetic editing ability,short generation time and fast developmental stage relatively.In this study,high-throughout sequencing was used to obtain a large number of transcriptome data on the trait strain of the stat1b gene,to study the stat1b gene effect on the molecular level.The zebrafish stat1b gene was knocked out using the CRISPR/Cas9gene editing technology and the stat1b gene mutant strain zebrafish was identified with DNA genome extraction,amplification of the target fragment,DNA recovery,T7 Endonuclease?digestion,TA clone and sanger sequencing,and succssfully obtainedthe stat1b F3 generation homozygous mutant strain zebrafish.In this article,the 1-cell period,2-cell period,4-cell period,8-cell period,and 16-cell period of the stat1b gene homozygous mutant and the wild-type embryo were respectively sequencing by the Illumina HiSeq platform.Differential genes and pathways were analyzed using the Gene Ontology?GO?enrichment and Kyoto Encyclopedia of Genes and Genomes?KEGG?pathways for each development stage embryo.The results showed there were 4791,3623,4319,7648,7889 genes was differentially screened in the embryo 1-cell period,2-cell period,4-cell period,8-cell period,and 16-cell period,respectively,compared with the wild-type zebrafish.The mutants were compared with the wild-type zebrafish embryos at the five cell stages.The top 50 differentially expressed genes were significantly up-regulated,the expression of fl1a gene was up-regulated,and the expression of adcy8and me3 genes was down-regulated The mutants were compared with the wild-type zebrafish embryos at the five cell stages.The top 50differentially expressed genes were significantly up-regulated,the expression of fl1a gene was up-regulated,and the expression of adcy8and me3 genes was down-regulated.GO enrichment analysis showed that the differential genes in the five cell periods were mainly enriched in metabolic processes,intracellular components,etc.The results of KEGG pathway enrichment analysis indicated that the differential genes in the five cell periods were mainly related to RNA transport and pan.Isolate-mediated proteolysis,protein processing on the endoplasmic reticulum and other pathways.According to the analysis of the result data,after the stat1b gene was knocked out,the expression of eif4e2,eif3jb and senp2 genes was down-regulated during the early development of zebrafish embryos,thereby affecting the RNA transport pathway.In addition,a preliminary screening of the differential expressed genes of the stat1b homozygous mutant strain and the wild-type zebrafish for each cell period was found to be associated with zinc finger proteins.It is speculated that the stat1b gene may be involved in cell differentiation,and revealed changes in the biological functions and pathways associated with stat1b gene mutations at the molecule Levels.