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Identification and expression analysis of microRNAs at the grain filling stage in rice(Oryza sativa L.)via deep sequencing.

Yi R, Zhu Z, Hu J, Qian Q, Dai J, Ding Y - PLoS ONE (2013)

Bottom Line: A total of 434 known miRNAs (380, 402, 390 and 392 at 5, 7, 12 and 17 days after fertilization, respectively.) were obtained from rice grain.In addition, sixty novel miRNAs were identified, and five of these were further validated experimentally.Additional analysis showed that the predicted targets of the differentially expressed miRNAs may participate in signal transduction, carbohydrate and nitrogen metabolism, the response to stimuli and epigenetic regulation.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Hybrid Rice, Department of Genetics, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.

ABSTRACT
MicroRNAs (miRNAs) have been shown to play crucial roles in the regulation of plant development. In this study, high-throughput RNA-sequencing technology was used to identify novel miRNAs, and to reveal miRNAs expression patterns at different developmental stages during rice (Oryza sativa L.) grain filling. A total of 434 known miRNAs (380, 402, 390 and 392 at 5, 7, 12 and 17 days after fertilization, respectively.) were obtained from rice grain. The expression profiles of these identified miRNAs were analyzed and the results showed that 161 known miRNAs were differentially expressed during grain development, a high proportion of which were up-regulated from 5 to 7 days after fertilization. In addition, sixty novel miRNAs were identified, and five of these were further validated experimentally. Additional analysis showed that the predicted targets of the differentially expressed miRNAs may participate in signal transduction, carbohydrate and nitrogen metabolism, the response to stimuli and epigenetic regulation. In this study, differences were revealed in the composition and expression profiles of miRNAs among individual developmental stages during the rice grain filling process, and miRNA editing events were also observed, analyzed and validated during this process. The results provide novel insight into the dynamic profiles of miRNAs in developing rice grain and contribute to the understanding of the regulatory roles of miRNAs in grain filling.

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Differential expression analysis of known miRNAs.(A) Heatmap for clustering analysis of the differentially expressed known miRNAs. The bar represents the scale of the expression levels of the miRNAs (log 2). (B) Validation via quantitative real-time RT-PCR of differentially expressed miRNAs obtained from deep sequencing. U6 snRNA was used as a reference, and the expression levels of each of the miRNAs were then compared with the expression at 5 DAF, which was set to 1.0. Error bars indicate the standard deviation (±SD) of three replicates.
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pone-0057863-g003: Differential expression analysis of known miRNAs.(A) Heatmap for clustering analysis of the differentially expressed known miRNAs. The bar represents the scale of the expression levels of the miRNAs (log 2). (B) Validation via quantitative real-time RT-PCR of differentially expressed miRNAs obtained from deep sequencing. U6 snRNA was used as a reference, and the expression levels of each of the miRNAs were then compared with the expression at 5 DAF, which was set to 1.0. Error bars indicate the standard deviation (±SD) of three replicates.

Mentions: Based on the normalized read count for each identified miRNA, differential expression analysis was performed, and 161 known rice miRNAs were found to show statistically significant (P<0.01) changes in their relative abundance in at least one of the three transitions (5 DAF to 7 DAF/7 DAF to 12 DAF/12 DAF to 17 DAF) during rice grain filling (Table S4), the majority of which are non-conserved miRNAs. A total of 161 known miRNAs were subjected to cluster analysis according to their expression profiles at different seed filling stages (Figure S2). The results showed that a large numbers of miRNAs were up-regulated during the 5 DAF to 7 DAF periods (cluster 20 in Fig. 3A). Some miRNAs were up-regulated gradually during grain filling (clusters 11, 13 and 18 in Fig. 3A), while some, in cluster 14, appeared to show a similar trend of down-regulation over the same period (Fig. 3A). Some of the miRNAs were preferentially expressed in at least one grain filling stage. For example, the miRNAs in clusters 2, 16 and 17 showed patterns that were enriched at 17 DAF, 7 DAF and 12 DAF, respectively, while the miRNAs in cluster 10 accumulated and peaked at 7 DAF, decreased greatly at 12 DAF, and were up-regulated thereafter (Fig. 3A).


Identification and expression analysis of microRNAs at the grain filling stage in rice(Oryza sativa L.)via deep sequencing.

Yi R, Zhu Z, Hu J, Qian Q, Dai J, Ding Y - PLoS ONE (2013)

Differential expression analysis of known miRNAs.(A) Heatmap for clustering analysis of the differentially expressed known miRNAs. The bar represents the scale of the expression levels of the miRNAs (log 2). (B) Validation via quantitative real-time RT-PCR of differentially expressed miRNAs obtained from deep sequencing. U6 snRNA was used as a reference, and the expression levels of each of the miRNAs were then compared with the expression at 5 DAF, which was set to 1.0. Error bars indicate the standard deviation (±SD) of three replicates.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585941&req=5

pone-0057863-g003: Differential expression analysis of known miRNAs.(A) Heatmap for clustering analysis of the differentially expressed known miRNAs. The bar represents the scale of the expression levels of the miRNAs (log 2). (B) Validation via quantitative real-time RT-PCR of differentially expressed miRNAs obtained from deep sequencing. U6 snRNA was used as a reference, and the expression levels of each of the miRNAs were then compared with the expression at 5 DAF, which was set to 1.0. Error bars indicate the standard deviation (±SD) of three replicates.
Mentions: Based on the normalized read count for each identified miRNA, differential expression analysis was performed, and 161 known rice miRNAs were found to show statistically significant (P<0.01) changes in their relative abundance in at least one of the three transitions (5 DAF to 7 DAF/7 DAF to 12 DAF/12 DAF to 17 DAF) during rice grain filling (Table S4), the majority of which are non-conserved miRNAs. A total of 161 known miRNAs were subjected to cluster analysis according to their expression profiles at different seed filling stages (Figure S2). The results showed that a large numbers of miRNAs were up-regulated during the 5 DAF to 7 DAF periods (cluster 20 in Fig. 3A). Some miRNAs were up-regulated gradually during grain filling (clusters 11, 13 and 18 in Fig. 3A), while some, in cluster 14, appeared to show a similar trend of down-regulation over the same period (Fig. 3A). Some of the miRNAs were preferentially expressed in at least one grain filling stage. For example, the miRNAs in clusters 2, 16 and 17 showed patterns that were enriched at 17 DAF, 7 DAF and 12 DAF, respectively, while the miRNAs in cluster 10 accumulated and peaked at 7 DAF, decreased greatly at 12 DAF, and were up-regulated thereafter (Fig. 3A).

Bottom Line: A total of 434 known miRNAs (380, 402, 390 and 392 at 5, 7, 12 and 17 days after fertilization, respectively.) were obtained from rice grain.In addition, sixty novel miRNAs were identified, and five of these were further validated experimentally.Additional analysis showed that the predicted targets of the differentially expressed miRNAs may participate in signal transduction, carbohydrate and nitrogen metabolism, the response to stimuli and epigenetic regulation.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Hybrid Rice, Department of Genetics, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.

ABSTRACT
MicroRNAs (miRNAs) have been shown to play crucial roles in the regulation of plant development. In this study, high-throughput RNA-sequencing technology was used to identify novel miRNAs, and to reveal miRNAs expression patterns at different developmental stages during rice (Oryza sativa L.) grain filling. A total of 434 known miRNAs (380, 402, 390 and 392 at 5, 7, 12 and 17 days after fertilization, respectively.) were obtained from rice grain. The expression profiles of these identified miRNAs were analyzed and the results showed that 161 known miRNAs were differentially expressed during grain development, a high proportion of which were up-regulated from 5 to 7 days after fertilization. In addition, sixty novel miRNAs were identified, and five of these were further validated experimentally. Additional analysis showed that the predicted targets of the differentially expressed miRNAs may participate in signal transduction, carbohydrate and nitrogen metabolism, the response to stimuli and epigenetic regulation. In this study, differences were revealed in the composition and expression profiles of miRNAs among individual developmental stages during the rice grain filling process, and miRNA editing events were also observed, analyzed and validated during this process. The results provide novel insight into the dynamic profiles of miRNAs in developing rice grain and contribute to the understanding of the regulatory roles of miRNAs in grain filling.

Show MeSH