SOX12基因在Vero细胞感染猪流行性腹泻病毒过程中的功能研究

doi:10.16431/j.cnki.1671-7236.2025.01.026

Abstract

Abstract: 【Objective】 The aim of this study was to investigate the effect of SOX12 gene on the replication of Porcine epidemic diarrhea virus (PEDV),so as to provide the effective molecular marker for anti-PEDV breeding. 【Method】 Three SOX12 gene interference sequences (siRNA1,siRNA2 and siRNA3) and a negative control sequence (siNC) were synthesized and transfected into African green monkey kidney cells (Vero cells) in this study.At 48 h after transfection,the cells were collected,and the interference efficiency was detected by Real-time quantitative PCR.Effective interference fragments and siNC were transfected into Vero cells for 24 h,respectively.Then,the cells were infected with PEDV at a multiplicity of infection of 0.05,and divided into four groups:PEDV-infected assay group at 12 h (12 hpi-siRNA),PEDV-infected control group at 12 h (12 hpi-siNC),PEDV-infected assay group at 24 h (24 hpi-siRNA),and PEDV-infected control group at 24 h (24 hpi-siNC).Cell samples from each group were collected and their RNA was extracted.The expression of PEDV N and SOX12 genes was detected by Real-time quantitative PCR,and the expression of PEDV N protein was analyzed by Western blotting.The viral titer of PEDV was determined through 50% tissue culture infective dose (TCID₅₀) assay,and the replication of PEDV of cells in each group was detected using indirect immunofluorescence (IFA) method.The SOX12 interaction genes were predicted using GeneMANIA website,and the expression changes of these interaction genes were detected by Real-time quantitative PCR after inhibiting SOX12 gene expression. 【Result】 The interference efficiency of the three SOX12 interfering fragments was 30% to 60%,with siRNA3 showing the highest efficiency of 60%.Compared with 12 hpi-siNC and 24 hpi-siNC groups,the transcription and protein expression of PEDV N gene in 12 hpi-siRNA and 24 hpi-siRNA groups was significantly decreased,respectively (P＜0.05).The results of the viral titer phenotype determination indicated that the viral titers of PEDV in 12 hpi-siRNA and 24 hpi-siRNA groups were significantly lower than that in their control groups,respectively (P＜0.05).IFA results also showed that PEDV replication in 12 hpi-siRNA and 24 hpi-siRNA groups was significantly less than that in control group.The gene interaction prediction and Real-time quantitative PCR detection results revealed that compared with siNC group,the expression of interacting genes (SOX17,DDX51,ZMIZ2,SSRP1,TAF6 and ABCB8) were significantly decreased after interfering with SOX12 gene expression (P＜0.05). 【Conclusion】 This study revealed that the regulatory role of SOX12 gene in the process of PEDV infection in Vero cells,and found that the downregulation of SOX12 gene expression significantly inhibited PEDV replication and the expression of its interacting genes SOX17, DDX51,ZMIZ2,SSRP1,TAF6 and ABCB8.

Key words: Porcine epidemic diarrhea virus (PEDV); SOX12 gene; Vero cells; gene interference

CLC Number:

S852.65⁺1

XIANG Jiaojiao, YUAN Na, LI Huihui, SHAO Mingzhu, ZHAO Fuping, ZHANG Longchao, WANG Lixian, SHI Lijun, CHEN Bin. Function Study on SOX12 Gene in Vero Cells Infected with Porcine Epidemic Diarrhea Virus[J]. China Animal Husbandry and Veterinary Medicine, 2025, 52(1): 289-297.

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Function Study on SOX12 Gene in Vero Cells Infected with Porcine Epidemic Diarrhea Virus

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