Screening and Identification of miRNA Regulating Porcine E-twenty-six Variant Gene 5

doi:10.13560/j.cnki.biotech.bull.1985.2021-1057

Abstract

Abstract:

E-twenty-six Variant Gene 5（ETV5）is a transcription factor that plays key role in the self-renewal and maintenance of spermatogonial stem cells. To screen the miRNAs regulating porcine ETV5 gene,the online analysis software TargetScan and MiRanda were used to predict the miRNAs that regulate ETV5,and dual luciferase activity detection was adapted to verify the screened conserved miRNAs. Then the verified miRNA mimics or inhibitor were transfected into porcine fetal fibroblasts and testicular Sertoli cells respectively,the expression of ETV5 was detected by real time fluorescence quantitative PCR. Concurrently,ETV5 gene was knocked out in the porcine fetal fibroblasts to detect the expression of miR-19. The results showed that miR-19 significantly reduced the activity of luciferase in the cells by binding to the 3`UTR region of ETV5（P<0.01）. Results of cell transfection demonstrated that miR-19 mimics significantly reduced the expression of ETV5 in the porcine fetal fibroblasts and testicular Sertoli cells（P<0.01）,while miR-19 inhibitor led to the opposite result（P<0.01）. After knockout of ETV5 gene in the fetal fibroblasts,the expression of the miR-19 significantly increased（P<0.01）. In sum,miR-19 may regulate the expression of porcine ETV5 gene,which provides a reference for further research on the effect of miRNA regulating porcine ETV5 gene on porcine reproduction.

Key words: miR-19, porcine, ETV5, testis

LI Hong-yi, PENG Guo-liang, XIAO Zheng-zhong, ZHANG Mao. Screening and Identification of miRNA Regulating Porcine E-twenty-six Variant Gene 5[J]. Biotechnology Bulletin, 2022, 38(5): 169-174.

Figures/Tables 5

Fig. 1 Prediction of miRNA regulating porcine ETV5 gene

Fig.2 Detection of luciferase activity ** indicate that there is a significant difference compared with the control group,P<0.01,the same below

Fig.3 MIR-19 regulating the mRNA expression of ETV5 in the fetal fibroblasts

Fig.4 MIR-19 regulating the mRNA expression of ETV5 in the Sertoli cells

Fig.5 Expression of miR-19 in the ETV5 knockout cells

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