miR-192在尼罗罗非鱼应答碱度胁迫中的表达及靶基因

采用实时荧光定量、生物信息学、双荧光素酶报告和体内miRNA抑制的方法，探究miR-192在尼罗罗非鱼（Oreochromis niloticus）应答碳酸盐碱度胁迫中的作用。结果如下：（1）实时荧光定量PCR实验表明，在急性碱度胁迫（6 g/L NaHCO₃）尼罗罗非鱼6 h后，鳃组织中miR-192的表达显著下调，溶质转运蛋白基因（solute carriers16A7，SLC16A7）表达显著上调（P<0.05）；（2）借助生物信息分析预测到SLC16A7可能是miR-192的靶基因；（3）利用双荧光素酶报告实验发现miR-192会与SLC16A7的3''UTR结合；（4）体内对miR-192抑制后，SLC16A7的表达显著上调（P<0.05）。证实了miR-192参与了尼罗罗非鱼应答碱度胁迫中的调控过程，SLC16A7是miR-192的直接靶基因，为探明miRNAs调控尼罗罗非鱼应答碱度胁迫的分子机制提供了一定的依据。

Expression of miR-192 in Nile tilapia in response to alkalinity stress and verification of target genes

In order to explore the role of miR-192 in response to carbonate alkalinity stress in Nile tilapia, real-time PCR, bioinformatics, dual-luciferase reporter assay and miRNA inhibition in vivo were used. The results were as follows:(1) The real-time PCR experiments showed that the expression of miR-192 was significantly down-regulated, while solute carriers16A7,SLC16A7 was significantly up-regulated in gills of Nile tilapia after 6 hours acute alkalinity stress (6 g/L NaHCO₃)(P<0.05); (2) Bioinformatics analysis revealed that SLC16A7 might be the target gene of miR-192; (3) Dual-luciferase reporter assay showed that miR-192 directly regulated SLC16A7 by targeting its 3''UTR; (4) Inhibition of miR-192 significantly increased the mRNA level of SLC16A7 in vivo (P<0.05). In conclusion, miR-192 was involved in the adaptation of Nile tilapia to alkalinity stress,and SLC16A7 was the direct target gene of miR-192. This study provides a basis for exploring the mechanism of miRNAs in the response to alkalinity stress in Nile tilapia.