Annals of Applied Biology ISSN 0003-4746 RESEARCH ARTICLE Hexaploid wheat (Triticum aestivum) root miRNome analysis in response to salt stress H. Eren 1,† , M.Y. Pekmezci 2,† , S. Okay 2 , M. Turktas 2 , B. Inal 2 , E. Ilhan 1 , M. Atak 3 , M. Erayman 1 & T. Unver 2 1 Department of Biology, Faculty of Science, Cankiri Karatekin University, Cankiri, Turkey 2 Department of Biology, Faculty of Arts and Science, Mustafa Kemal University, Hatay, Turkey 3 Department of Field Crops, Faculty of Agriculture, Mustafa Kemal University, Hatay, Turkey Keywords Microarray; miRNA; salt stress; transcriptome; wheat. Correspondence T. Unver, Department of Biology, Faculty of Science, Cankiri Karatekin University, 18100 Cankiri, Turkey. Email: turgayunver@gmail.com † These authors contributed equally to this study. Received: 29 July 2014; revised version accepted: 16 March 2015; published online: 13 May 2015. doi:10.1111/aab.12219 Abstract Hexaploid bread wheat (Triticum aestivum) is one of the major crops grown and consumed all over the world. Elevated soil salinity causes reduction in crop yield and quality; therefore, several strategies were developed to improve salt-tolerant cultivars. MicroRNAs (miRNAs), small and non-coding RNAs, regulate gene expression at post-transcriptional level and play important roles in stress tolerance. Here, we used a broad-range miRNA-microarray analysis to investigate the root-miRNA profiles of two cultivars, Bezostaja (sensitive) and Seri-82 (tolerant). A total of 44 differentially regulated miRNAs were identified in the 8 × 15K array containing 11 862 plant miRNAs available in the database. Sixteen novel salt-responsive miRNAs were determined in wheat for the first time. The expression of three miRNAs (hvu-miR5049a, ppt-miR1074 and osa-miR444b.2) was up-regulated more than 260-fold in cv. Bezostaja upon salt stress. The target-gene analyses showed that sev- eral salt-stress-responsive miRNAs regulate mainly transcription factors such as bHLH135-like, AP2/ERBP, MADS-box and transporters. Gene ontol- ogy searches for 565 putative salt-stress-responsive miRNA target-genes revealed 623 processes in 10 different main topics such as metabolic process and response to stimuli. The genome-wide root miRNome study indicates salt-stress-responsive wheat miRNAs and the possible mechanisms behind the tolerance. Introduction Wheat (Triticum aestivum L.) is one of the staple crops being an essential food source covering 18.8% of daily calorie intake in human diet; its global production is about 670.8 million tonnes per annum in 215.4 million ha of land (FAO, 2012). Wheat is harvested all over the world for production of various foods, such as bread, pasta, noodles, cakes and biscuits (Shewry & Tatham, 1997). Plants encounter many biotic and abiotic stress factors resulting in various levels of yield loss (Okay et al., 2014). Salt stress is one of the major abiotic stress factors affecting photosynthesis, growth and development in plants (Nemoto & Sasakuma, 2002). The microRNAs (miRNAs) are a class of small (21–24 nucleotides in length), endogenously expressed and non-coding RNA molecules. The miRNA genes are expressed as primary miRNAs (pri-miRNAs) and pro- cessed into pre-miRNAs by a dicer-like enzyme (DCL1). The pre-miRNAs are methylated and finally form mature miRNAs which are incorporated into RNA-induced silencing complex (RISC). The miRNA–RISC regulates gene expression via post-transcriptional inhibition and cleavage (Eldem et al., 2013). The regulatory role of plant miRNAs was first described by Reinhart et al. (2002) in Arabidopsis. Involvement of miRNAs in tolerance to vari- ous stress conditions as well as growth and development has been shown in several plant species (Jung & Park, 2007; Trindade et al., 2010; Xin et al., 2010; Fu et al., 2012). To date, a number of wheat miRNAs have been identified by several approaches. In a recent study, a 208 Ann Appl Biol 167 (2015) 208–216 © 2015 Association of Applied Biologists