Identification of miRNAs and their targets in tomato at moderately and acutely elevated temperatures by high-throughput sequencing and degradome analysis

MicroRNAs (miRNAs) are 19-24 nucleotide (nt) noncoding RNAs that play important roles in abiotic stress responses in plants. High temperatures have been the subject of considerable attention due to their negative effects on plant growth and development. Heat-responsive miRNAs have been identified in some plants. However, there have been no reports on the global identification of miRNAs and their targets in tomato at high temperatures, especially at different elevated temperatures. Here, three small-RNA libraries and three degradome libraries were constructed from the leaves of the heat-tolerant tomato at normal, moderately and acutely elevated temperatures (26/18°C, 33/33°C and 40/40°C, respectively). Following high-throughput sequencing, 662 conserved and 97 novel miRNAs were identified. Of these miRNAs, 96 and 150 miRNAs were responsive to the moderately and acutely elevated temperature, respectively. Following degradome sequencing, 349 sequences were identified as targets of 138 conserved miRNAs, and 13 sequences were identified as targets of eight novel miRNAs. The expression levels of four miRNAs and five target genes obtained by quantitative real-time PCR (qRT-PCR) were largely consistent with the sequencing results. This study enriches the number of heat-responsive miRNAs and lays a foundation for the elucidation of the miRNA-mediated regulatory mechanism in tomatoes at elevated temperatures. Overall design: The seedlings were randomly divided into three groups for temperature treatments on the seventh day after transfer. The first group was kept at a normal temperature and used as a control (26/18°C, NT); the second group was exposed to a moderately elevated temperature (33/33°C, MET); and the third group was exposed to an acutely elevated temperature (40/40°C, AET). The temperature reached 33°C in half an hour and 40°C in an hour in the climate chamber. All of them was used to do examination of small RNA and degradome RNA.