ORIGINAL PAPER New wheat microRNA using whole-genome sequence Kuaybe Yucebilgili Kurtoglu & Melda Kantar & Hikmet Budak Received: 16 September 2013 /Revised: 6 December 2013 /Accepted: 22 December 2013 /Published online: 7 January 2014 # Springer-Verlag Berlin Heidelberg 2014 Abstract MicroRNAs are post-transcriptional regulators of gene expression, taking roles in a variety of fundamental biological processes. Hence, their identification, annotation and characterization are of great significance, especially in bread wheat, one of the main food sources for humans. The recent availability of 5× coverage Triticum aestivum L. whole- genome sequence provided us with the opportunity to perform a systematic prediction of a complete catalogue of wheat microRNAs. Using an in silico homology-based approach, stem-loop coding regions were derived from two assemblies, constructed from wheat 454 reads. To avoid the presence of pseudo-microRNAs in the final data set, transposable element related stem-loops were eliminated by repeat analysis. Over- all, 52 putative wheat microRNAs were predicted, including seven, which have not been previously published. Moreover, with distinct analysis of the two different assemblies, both variety and representation of putative microRNA-coding stem-loops were found to be predominant in the intergenic regions. By searching available expressed sequences and small RNA library databases, expression evidence for 39 (out of 52) putative wheat microRNAs was provided. Expres- sion of three of the predicted microRNAs (miR166, miR396 and miR528) was also comparatively quantified with real- time quantitative reverse transcription PCR. This is the first report on in silico prediction of a whole repertoire of bread wheat microRNAs, supported by the wet-lab validation. Keywords Triticum aestivum . MicroRNA . MicroRNA prediction . Next-generation sequencing . Real-time quantitative reverse transcription PCR Abbreviations miRNA MicroRNA pri-miRNA Primary miRNA pre-miRNA Precursor miRNA RISC RNA-induced silencing complex SVM Support vector machine EST Expressed sequence tag LCG Low copy number assembly TREP Triticeae repeat database OG Orthologous group assembly OGR Orthologous group representatives cDNA Complementary DNA MFE Minimal folding free energy MITE Miniature inverted terminal repeat element TE-MIR Transposable element-related miRNA group GEO Gene expression omnibus TIR Terminal inverted repeat LTR Long terminal repeat siRNA Small interfering RNA Introduction Bread wheat (Triticum aestivum L.) is one of the most exten- sively grown crops with a global annual production of over 650 million tones. It is a fundamental source for human food consumption, providing approximately 20 % of the dietary energy supply (http://www.fao.org, 2011). Hexaploid bread wheat genome is large (~17Gbp), highly repetitive (80 %) and complex, having three homeologous but divergent subgenomes (AABBDD, 2n=42) (Hernandez et al. 2012; Kuaybe Yucebilgili Kurtoglu and Melda Kantar equally contributed to this research. Electronic supplementary material The online version of this article (doi:10.1007/s10142-013-0357-9) contains supplementary material, which is available to authorized users. K. Y. Kurtoglu : M. Kantar : H. Budak (*) Biological Sciences and Bioengineering Program, Sabanci University, Orhanli 34956, Istanbul, Turkey e-mail: budak@sabanciuniv.edu Funct Integr Genomics (2014) 14:363–379 DOI 10.1007/s10142-013-0357-9