Zhou Rong, Wang Qian, Jiang Fangling, Cao Xue, Sun Mintao, Liu Min, Wu Zhen
College of Horticulture, Nanjing Agricultural University, Weigang No. 1, Nanjing, 210095, China.
Vegetable Research Institute, Jiangsu Academy of Agricultural Science, Zhongling Street No. 50, Nanjing, 210014, China.
Sci Rep. 2016 Sep 22;6:33777. doi: 10.1038/srep33777.
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 in total with 469 conserved and 91 novel miRNAs shared in the three small-RNA libraries. 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 seven miRNAs and six 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.
微小RNA（miRNA）是19 - 24个核苷酸（nt）的非编码RNA，在植物非生物胁迫反应中发挥重要作用。高温因其对植物生长发育的负面影响而受到广泛关注。在一些植物中已鉴定出热响应性miRNA。然而，关于番茄在高温下，尤其是在不同升高温度下miRNA及其靶标的全面鉴定尚无报道。在此，从耐热番茄叶片分别在正常、中度和急性升高温度（分别为26/18°C、33/33°C和40/40°C）下构建了三个小RNA文库和三个降解组文库。经过高通量测序，共鉴定出662个保守miRNA和97个新miRNA，其中469个保守miRNA和91个新miRNA在三个小RNA文库中共享。在这些miRNA中，分别有96个和150个miRNA对中度和急性升高温度有响应。经过降解组测序，349个序列被鉴定为138个保守miRNA的靶标，13个序列被鉴定为8个新miRNA的靶标。通过定量实时PCR（qRT-PCR）获得的7个miRNA和6个靶基因的表达水平与测序结果基本一致。本研究丰富了热响应性miRNA的数量，为阐明番茄在高温下miRNA介导的调控机制奠定了基础。
