Characterization of Expression Patterns of Grapevine MicroRNA Family Members using MicroRNA Rapid Amplification of Complementary DNA Ends.

Abstract

Grapevine (Vitis vinifera L.), with important nutritional values and health benefits, is one of the most economically fruit crop worldwide. In the present study, real-time quantitative polymerase chain reaction (qRT-PCR) and microRNA rapid amplification of cDNA ends (miR-RACE) techniques were used to characterize the expression and diversification patterns of various grapevine microRNAs (Vv-miRNAs) and their family members in grapevine. Based on our results, eight different grapevine miRNAs (miR159, miR164, miR167, miR172, miR319, miR393, miR396, and miR398) and their family members were expressed in different tissues at various developmental stages. The qRT-PCR results showed that the expression levels of Vv-miRNAs during grapevine development were dynamic. Furthermore, based on miR-RACE analysis and polymerase chain reaction (PCR) product sequencing results, different members within the same miRNA family were also expressed at different levels. Comparing the spatiotemporal expression levels of different members in the same miRNA family indicated that some miRNA families might have a key miRNA member that played the prominent role in regulation of their subsequent common target genes. In conclusion, our results showed that miR-RACE is a powerful technique to analyze the expression patterns of different members in the same miRNA family in terms of reverse-transcription (RT) efficiency and specificity. The findings of the expression diversification among Vv-miRNA family members and the existence of some Vv-miRNAs playing the key role could add to our understanding about the regulatory role of miRNAs in grapevine.