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      Deep mRNA sequencing reveals stage-specific transcriptome alterations during microsclerotia development in the smoke tree vascular wilt pathogen, Verticillium dahliae

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          Abstract

          Background

          Verticillium dahliae is a soil-borne fungus that causes vascular wilt diseases in a wide range of plant hosts. V. dahliae produces multicelled, melanized resting bodies, also known as microsclerotia (MS) that can survive for years in the soil. The MS are the primary source of infection of the Verticillium disease cycle. Thus, MS formation marks an important event in the disease cycle of V. dahliae.

          Results

          In this study, next generation sequencing technology of RNA-Seq was employed to investigate the global transcriptomic dynamics of MS development to identify differential gene expression at several stages of MS formation in strain XS11 of V. dahliae, isolated from smoke tree. We observed large-scale changes in gene expression during MS formation, such as increased expression of genes involved in protein metabolism and carbohydrate metabolism. Genes involved in glycolytic pathway and melanin biosynthesis were dramatically up-regulated in MS. Cluster analyses revealed increased expression of genes encoding products involved in primary metabolism and stress responses throughout MS development. Differential expression of ubiquitin-dependent protein catabolism and cell death-associated genes during MS development were revealed. Homologs of genes located in the lineage-specific (LS) regions of V. dahliae strain VdLs.17, were either not expressed or showed low expression. Furthermore, alternative splicing (AS) events were analyzed, revealing that over 95.0% AS events involve retention of introns (RI).

          Conclusions

          These data reveal the dynamics of transcriptional regulation during MS formation and were used to construct a comprehensive high-resolution gene expression map. This map provides a key resource for understanding the biology and molecular basis of MS development of V. dahliae.

          Electronic supplementary material

          The online version of this article (doi:10.1186/1471-2164-15-324) contains supplementary material, which is available to authorized users.

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          Most cited references57

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          The transcriptional landscape of the yeast genome defined by RNA sequencing.

          The identification of untranslated regions, introns, and coding regions within an organism remains challenging. We developed a quantitative sequencing-based method called RNA-Seq for mapping transcribed regions, in which complementary DNA fragments are subjected to high-throughput sequencing and mapped to the genome. We applied RNA-Seq to generate a high-resolution transcriptome map of the yeast genome and demonstrated that most (74.5%) of the nonrepetitive sequence of the yeast genome is transcribed. We confirmed many known and predicted introns and demonstrated that others are not actively used. Alternative initiation codons and upstream open reading frames also were identified for many yeast genes. We also found unexpected 3'-end heterogeneity and the presence of many overlapping genes. These results indicate that the yeast transcriptome is more complex than previously appreciated.
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            The impact of next-generation sequencing technology on genetics.

            If one accepts that the fundamental pursuit of genetics is to determine the genotypes that explain phenotypes, the meteoric increase of DNA sequence information applied toward that pursuit has nowhere to go but up. The recent introduction of instruments capable of producing millions of DNA sequence reads in a single run is rapidly changing the landscape of genetics, providing the ability to answer questions with heretofore unimaginable speed. These technologies will provide an inexpensive, genome-wide sequence readout as an endpoint to applications ranging from chromatin immunoprecipitation, mutation mapping and polymorphism discovery to noncoding RNA discovery. Here I survey next-generation sequencing technologies and consider how they can provide a more complete picture of how the genome shapes the organism.
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              Computational methods for transcriptome annotation and quantification using RNA-seq.

              High-throughput RNA sequencing (RNA-seq) promises a comprehensive picture of the transcriptome, allowing for the complete annotation and quantification of all genes and their isoforms across samples. Realizing this promise requires increasingly complex computational methods. These computational challenges fall into three main categories: (i) read mapping, (ii) transcriptome reconstruction and (iii) expression quantification. Here we explain the major conceptual and practical challenges, and the general classes of solutions for each category. Finally, we highlight the interdependence between these categories and discuss the benefits for different biological applications.
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                Author and article information

                Contributors
                xiongdianguang@126.com
                ylwang@bjfu.edu.cn
                majie@bjfu.edu.cn
                Steve.Klosterman@ars.usda.gov
                jessiecain4ever@163.com
                chengmt@bjfu.edu.cn
                Journal
                BMC Genomics
                BMC Genomics
                BMC Genomics
                BioMed Central (London )
                1471-2164
                1 May 2014
                1 May 2014
                2014
                : 15
                : 1
                : 324
                Affiliations
                [ ]The Key Laboratory for Silviculture and Conservation of Ministry of Education, College of Forestry, Beijing Forestry University, Beijing, China
                [ ]School of Information Science and Technology, Beijing Forestry University, Beijing, China
                [ ]United States Department of Agriculture-Agricultural Research Service, Salinas, CA USA
                Article
                6023
                10.1186/1471-2164-15-324
                4035056
                24884698
                dca7f34e-63cb-414e-8772-13a5e2e21493
                © Xiong et al.; licensee BioMed Central Ltd. 2014

                This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited.

                History
                : 27 August 2013
                : 22 April 2014
                Categories
                Research Article
                Custom metadata
                © The Author(s) 2014

                Genetics
                verticillium dahliae,microsclerotia development,rna-seq,transcriptome,gene expression,alternative splicing

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