动物造模 z-bio 2021-11-10 309

　　Objective: To investigate the regulatory effects of nuclear transcription factor TRB, RXR and agonist T4 on Cip4 gene expression in Mongolian gerbils.

　　Methods: Synthesize the sequence of the promoter region of the Cip4 gene of Mongolian gerbils, construct the reporter vector pGL3-Cip4; clone the transcription factor TRB and RXR into the pcDNA3.1 expression vector. PGL3-Cip4, pcDNA3.1-TRB, pcDNA3.1 -RXR and pRL-TK (reference plasmid) vectors were co-transfected into HEK-293 cells to construct a luciferase double reporter system for Cip4 gene. Observe the amount of plasmid and the effect of treatment with TRB agonist Thyroxine T4 on transcriptional activity.

　　Results: The dual luciferase reporter gene system with the Cip4 promoter region of Mongolian gerbils as the promoter sequence has the highest transfection efficiency when (promoter + transcription factor): pRL-TK=20:1 and the total plasmid amount is 2μg. It also has the highest luciferase activity. The test results show that the addition of TRB alone does not change the transcription level of Cip4 (P>0.05), and the addition of RXR alone can increase the transcription level of Cip4 (P<0.05). The combined effect of RXR and T4 can be upregulated The transcription level of Cip4 (P<0.001). The combination of TRB and T4 can down-regulate the activity of Cip4 (P<0.05). When RXR, TRB, and T4 coexist, the transcription level of Cip4 is significantly down-regulated (P<0.01).

　　Conclusion: This study confirms that T4, TRB and RXR co-regulate the transcription of the Cip4 gene in unguiculatus gerbils, mimicking the activation of the ligand-activated transcription factor TRB/RXR to activate the transcription of the target gene Cip4, and also confirms the transcription activation of Cip4 and the RXR signal The pathways are closely related, laying the foundation for revealing the mechanism of thyroxine in human NAFLD. This reporter gene system can be used for the analysis of the mechanism of ligand-activated transcription factor regulating Cip4 gene expression and the screening of related drugs.