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Validation of miRNAs targets through luciferase assay in bovine granulosa cells


MicroRNAs (miRNAs) are small noncoding RNAs involved in modulate mRNA levels by repression or degradation. MiRNAs are found in body fluids enclosed in extracellular vesicles. Recently, miRNAs studies demonstrated the importance of these molecules regulating target genes and biological pathways. Besides, extracellular vesicles containing miRNAs are potential biomarkers and tools for different treatments. Therefore, miRNAs studies can provide a better understand of pathologic and physiologic processes. Among them are the reproductive processes. In this scenario, miRNAs within the follicular environment are capable to modulate biologic mechanisms related to follicular development and oocyte maturation. Additionally, several studies have demonstrated the importance of communication between granulosa cells, cumulus cells and oocyte regarding follicular development and oocyte quality. This communication involves molecular transport as well as pathways modulation, which can be regulated by miRNAs. Additionally, the in silico analyses of miRNAs within the follicular environment can possible identify molecules and candidate pathways associated with follicle development and oocyte quality. These studies are able to identify potential miRNAs functions, but they do not allow the proof of their action towards the regulation of target genes. Thus, the luciferase assay technique is an important tool to validate the binding of miRNAs to the 3'UTR of target genes. The application of this technique has great impact in the development of functional studies using miRNAs, because it guarantees the binding of miRNAs to specific targets. Therefore, the use of this technique allows greater credibility to the studies. In conclusion, miRNAs are important targets for studies in reproduction processes. However, the interpretation of its functions remains broad, demonstrating the necessity to use refined techniques, such as the luciferase assay in order to develop precise functional studies. (AU)

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