Abstract：MiRNA is a non-coding small RNA in eukaryotic organisms. It plays an important role in the development of animals and plants. However, the molecular mechanism of rice resistance to high temperature participated by miRNA has seldom been studied. As an important staple food, rice (Oryza sativa) frequently suffers from heat stress during the growing season. Clarifying plant heat stress response and tolerance mechanism at miRNA level is the basis to breed varieties with a suitable heat stress (HS) tolerance. In previous study, six libraries (double for 0, 12 and 24 h heat treatment) constructed from heat tolerant variety Oryza sativa ssp. indica cv. HT54, were sequenced by next-generation sequencing (NGS) technique. Those study found that the miR396 family member and their putative target genes might play pivotal roles as regulator reacting to heat stress. In this study, the expression of all 8 miR396 family members and 12 predicted target genes were detected in seedlings of Oryza sativa spp. japonica cv. Nipponbare with 48 ℃ heat stress for 6 different time points. The results indicated that all 8 miR396 family members and 12 predicted target genes showed different reaction to heat stress. The expression level of miR396a, miR396e and miR396f increased at high temperature stress at each time points, the expression level increased up to 10 fold (in miR396a and miR396e), even 100-fold (in miR396f); while miR396b, miR396c, miR396d, miR396g and miR396h decreased at 1.5, 3, 6 h, but increased at 12, 24 h. Correspondingly, the OsGRF family showed irregular upward or descending expression with high temperature stress. The target gene OsGRF2 decreased up to 8~10-fold. OsGRF2 had a high negative correlation with the expression of miR396a, miR396e and miR396f, indicating that OsGRF2 was the target gene for miR396a, miR396e and miR396f. These results suggest that miR396 would play a key role in rice heat tolerance.
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