Dataset Information

Identification and characterization of microRNAs from barley (Hordeum vulgare L.) by high-throughput sequencing.

ABSTRACT: MicroRNAs (miRNAs) are a class of endogenous RNAs that regulates the gene expression involved in various biological and metabolic processes. Barley is one of the most important cereal crops worldwide and is a model organism for genetic and genomic studies in Triticeae species. However, the miRNA research in barley has lagged behind other model species in grass family. To obtain more information of miRNA genes in barley, we sequenced a small RNA library created from a pool of equal amounts of RNA from four different tissues using Solexa sequencing. In addition to 126 conserved miRNAs (58 families), 133 novel miRNAs belonging to 50 families were identified from this sequence data set. The miRNA* sequences of 15 novel miRNAs were also discovered, suggesting the additional evidence for existence of these miRNAs. qRT-PCR was used to examine the expression pattern of six randomly selected miRNAs. Some miRNAs involved in drought and salt stress response were also identified. Furthermore, the potential targets of these putative miRNAs were predicted using the psRNATarget tools. Our results significantly increased the number of novel miRNAs in barley, which should be useful for further investigation into the biological functions and evolution of miRNAs in barley and other species.

SUBMITTER: Lv S

PROVIDER: S-EPMC3317698 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Identification and characterization of microRNAs from barley (Hordeum vulgare L.) by high-throughput sequencing.

Lv Shuzuo S Nie Xiaojun X Wang Le L Du Xianghong X Biradar Siddanagouda S SS Jia Xiaoou X Weining Song S

International journal of molecular sciences 20120306 3

MicroRNAs (miRNAs) are a class of endogenous RNAs that regulates the gene expression involved in various biological and metabolic processes. Barley is one of the most important cereal crops worldwide and is a model organism for genetic and genomic studies in Triticeae species. However, the miRNA research in barley has lagged behind other model species in grass family. To obtain more information of miRNA genes in barley, we sequenced a small RNA library created from a pool of equal amounts of RNA ...[more]

PMID: 22489137

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Project description:Small RNAs (21-24 nt) are pivotal regulators of gene expression that guide both transcriptional and post-transcriptional silencing mechanisms in diverse eukaryotes, including most if not all plants. MicroRNAs (miRNAs) and short interfering RNAs (siRNAs) are the two major types, both of which have a demonstrated and important role in plant development, stress responses and pathogen resistance. In this work, we used a deep sequencing approach (Sequencing-By-Synthesis, or SBS) to develop sequence resources of small RNAs from different Hordeum vulgare tissues (leaves, inflorescence and leaves inoculated with Blumeria). The high depth of the resulting datasets enabled us to examine in detail critical small RNA features as size distribution, tissue-specific regulation and sequence conservation between different organs in this species. We also developed database resources and a dedicated website (http://smallrna.udel.edu/) with computational tools for allowing other users to identify new miRNAs or siRNAs involved in specific regulatory pathways, verify the degree of conservation of these sequences in other plant species and map small RNAs on genes or larger regions of the maize genome under study. Small RNA libraries were derived from leaves, inflorescence and leaves inoculated with Blumeria of Hordeum vulgare. Each tissue represented a mixture of developmental stages. Total RNA was isolated using the Plant RNA Purification Reagent (Invitrogen) and submitted to Illumina (Hayward, CA, http://www.illumina.com) for small RNA library construction using approaches described in (Lu et al., 2007) with minor modifications. The small RNA libraries were sequenced with the Sequencing-By-Synthesis (SBS) technology by Illumina. PERL scripts were designed to remove the adapter sequences and determine the abundance of each distinct small RNA. We thank Roger Wise for providing the plant material and Kan Nobuta and Gayathri Mahalingam for assistance with the computational methods..

Project description:BackgroundMany plant species have been investigated in the last years for the identification and characterization of the corresponding miRNAs, nevertheless extensive studies are not yet available on barley (at the time of this writing). To extend and to update information on miRNAs and their targets in barley and to identify candidate polymorphisms at miRNA target sites, the features of previously known plant miRNAs have been used to systematically search for barley miRNA homologues and targets in the publicly available ESTs database. Matching sequences have then been related to Unigene clusters on which most of this study was based.ResultsOne hundred-fifty-six microRNA mature sequences belonging to 50 miRNA families have been found to significantly match at least one EST sequence in barley. As expected on the basis of phylogenetic relations, miRNAs putatively orthologous to those of Triticum are significantly over-represented inside the set of identified barley microRNA mature sequences. Many previously known and several putatively new miRNA/target pairs have been identified. When the predicted microRNA targets were grouped into functional categories, biological processes previously known to be regulated by miRNAs, such as development and response to biotic and abiotic stress, have been highlighted and most of the target molecular functions were related to transcription regulation. Candidate microRNA coding genes have been reported and genetic variation (SNPs/indels) both in functional regions of putative miRNAs (mature sequence) and at miRNA target sites has been found.ConclusionsThis study has provided an update of the information on barley miRNAs and their targets representing a foundation for future studies. Many of previously known plant microRNAs have homologues in barley with expected important roles during development, nutrient deprivation, biotic and abiotic stress response and other important physiological processes. Putative polymorphisms at miRNA target sites have been identified and they can represent an interesting source for the identification of functional genetic variability.

Dataset Information

Identification and characterization of microRNAs from barley (Hordeum vulgare L.) by high-throughput sequencing.

Publications

Identification and characterization of microRNAs from barley (Hordeum vulgare L.) by high-throughput sequencing.

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