Sorghum is largely grown for food, fodder and for biofuel production in semi-arid regions where the drought or high temperature or their combination co-occur. Plant microRNAs (miRNAs) are integral to the gene regulatory networks that control almost all biological processes including adaptation to stress conditions. Thus far, plant miRNA profiles under separate drought or heat stresses have been reported but not under combined drought and heat. In this study, we report miRNA profiles in leaves of sorghum exposed to individual drought or heat or their combination. Approximately 29 conserved miRNA families represented by 80 individual miRNAs, 26 families represented by 47 members of less conserved or sorghum-specific miRNA families as well as 8 novel miRNA families have been identified. Of these, 25 miRNAs were found to be differentially regulated in response to stress treatments. The comparative profiling revealed that the miRNA regulation was stronger under heat or combination of heat and drought compared to the drought alone. Furthermore, using degradome sequencing, 48 genes were confirmed as targets for the miRNAs in sorghum. Overall, this study provides a framework for understanding of the miRNA-guided gene regulations under combined stresses.
In plants, microRNAs (miRNAs) regulate their mRNA targets by precisely guiding cleavages between the 10th and 11th nucleotides in the complementary regions. High-throughput sequencing-based methods, such as PARE or degradome profiling coupled with a computational analysis of the sequencing data, have recently been developed for identifying miRNA targets on a genome-wide scale. The existing algorithms limit the number of mismatches between a miRNA and its targets and strictly do not allow a mismatch or G:U Wobble pair at the position 10 or 11. However, evidences from recent studies suggest that cleavable targets with more mismatches exist indicating that a relaxed criterion can find additional miRNA targets. In order to identify targets including the ones with weak complementarities from degradome data, we developed a computational method called SeqTar that allows more mismatches and critically mismatch or G:U pair at the position 10 or 11. Precisely, two statistics were introduced in SeqTar, one to measure the alignment between miRNA and its target and the other to quantify the abundance of reads at the center of the miRNA complementary site. By applying SeqTar to publicly available degradome data sets from Arabidopsis and rice, we identified a substantial number of novel targets for conserved and non-conserved miRNAs in addition to the reported ones. Furthermore, using RLM 5'-RACE assay, we experimentally verified 12 of the novel miRNA targets (6 each in Arabidopsis and rice), of which some have more than 4 mismatches and have mismatches or G:U pairs at the position 10 or 11 in the miRNA complementary sites. Thus, SeqTar is an effective method for identifying miRNA targets in plants using degradome data sets. 相似文献
In this article, a novel, label-free, and inherent electroactive redox immunosensor for carcinoembryonic antigen (CEA) based on gold nanoparticles (AuNPs) and Nile blue A (NB) hybridized electrochemically reduced graphene oxide (NB–ERGO) is proposed. The composite of NB–graphene oxide (NB–GO) was prepared by π–π stacking interaction. Then, chronoamperometry was adopted to simultaneously reduce HAuCl4 and nanocomposites of NB–GO for synthesizing AuNPs/NB–ERGO. The immunosensor was fabricated by capturing CEA antibody (anti-CEA) at this nanocomposite modified electrode. The immunosensor determination was based on the fact that, due to the formation of antigen–antibody immunocomplex, the decreased response currents of NB were directly proportional to the concentrations of CEA. Under optimal conditions, the linear range of the proposed immunosensor was estimated to be from 0.001 to 40 ng ml−1 and the detection limit was estimated to be 0.00045 ng ml−1. The proposed immunosensor was used to determine CEA in clinical serum samples with satisfactory results. The proposed method may provide promising potential application in clinical immunoassays with the properties of facile procedure, stability, high sensitivity, and selectivity. 相似文献