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High Quality Unigenes and Microsatellite Markers from Tissue Specific Transcriptome and Development of a Database in Clusterbean (Cyamopsis tetragonoloba, L. Taub)

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      Abstract

      Clusterbean (Cyamopsis tetragonoloba L. Taub), is an important industrial, vegetable and forage crop. This crop owes its commercial importance to the presence of guar gum (galactomannans) in its endosperm which is used as a lubricant in a range of industries. Despite its relevance to agriculture and industry, genomic resources available in this crop are limited. Therefore, the present study was undertaken to generate RNA-Seq based transcriptome from leaf, shoot, and flower tissues. A total of 145 million high quality Illumina reads were assembled using Trinity into 127,706 transcripts and 48,007 non-redundant high quality (HQ) unigenes. We annotated 79% unigenes against Plant Genes from the National Center for Biotechnology Information (NCBI), Swiss-Prot, Pfam, gene ontology (GO) and KEGG databases. Among the annotated unigenes, 30,020 were assigned with 116,964 GO terms, 9984 with EC and 6111 with 137 KEGG pathways. At different fragments per kilobase of transcript per millions fragments sequenced (FPKM) levels, genes were found expressed higher in flower tissue followed by shoot and leaf. Additionally, we identified 8687 potential simple sequence repeats (SSRs) with an average frequency of one SSR per 8.75 kb. A total of 28 amplified SSRs in 21 clusterbean genotypes resulted in polymorphism in 13 markers with average polymorphic information content (PIC) of 0.21. We also constructed a database named ‘ClustergeneDB’ for easy retrieval of unigenes and the microsatellite markers. The tissue specific genes identified and the molecular marker resources developed in this study is expected to aid in genetic improvement of clusterbean for its end use.

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      Basic local alignment search tool.

      A new approach to rapid sequence comparison, basic local alignment search tool (BLAST), directly approximates alignments that optimize a measure of local similarity, the maximal segment pair (MSP) score. Recent mathematical results on the stochastic properties of MSP scores allow an analysis of the performance of this method as well as the statistical significance of alignments it generates. The basic algorithm is simple and robust; it can be implemented in a number of ways and applied in a variety of contexts including straightforward DNA and protein sequence database searches, motif searches, gene identification searches, and in the analysis of multiple regions of similarity in long DNA sequences. In addition to its flexibility and tractability to mathematical analysis, BLAST is an order of magnitude faster than existing sequence comparison tools of comparable sensitivity.
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        edgeR: a Bioconductor package for differential expression analysis of digital gene expression data

        Summary: It is expected that emerging digital gene expression (DGE) technologies will overtake microarray technologies in the near future for many functional genomics applications. One of the fundamental data analysis tasks, especially for gene expression studies, involves determining whether there is evidence that counts for a transcript or exon are significantly different across experimental conditions. edgeR is a Bioconductor software package for examining differential expression of replicated count data. An overdispersed Poisson model is used to account for both biological and technical variability. Empirical Bayes methods are used to moderate the degree of overdispersion across transcripts, improving the reliability of inference. The methodology can be used even with the most minimal levels of replication, provided at least one phenotype or experimental condition is replicated. The software may have other applications beyond sequencing data, such as proteome peptide count data. Availability: The package is freely available under the LGPL licence from the Bioconductor web site (http://bioconductor.org). Contact: mrobinson@wehi.edu.au
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          Trimmomatic: a flexible trimmer for Illumina sequence data

          Motivation: Although many next-generation sequencing (NGS) read preprocessing tools already existed, we could not find any tool or combination of tools that met our requirements in terms of flexibility, correct handling of paired-end data and high performance. We have developed Trimmomatic as a more flexible and efficient preprocessing tool, which could correctly handle paired-end data. Results: The value of NGS read preprocessing is demonstrated for both reference-based and reference-free tasks. Trimmomatic is shown to produce output that is at least competitive with, and in many cases superior to, that produced by other tools, in all scenarios tested. Availability and implementation: Trimmomatic is licensed under GPL V3. It is cross-platform (Java 1.5+ required) and available at http://www.usadellab.org/cms/index.php?page=trimmomatic Contact: usadel@bio1.rwth-aachen.de Supplementary information: Supplementary data are available at Bioinformatics online.
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            Author and article information

            Affiliations
            [1 ]ICAR-National Research Centre on Plant Biotechnology, New Delhi 110012, India; hukam.rawal@ 123456gmail.com (H.C.R.); kumarshrawan12@ 123456gmail.com (S.K.); amithamithra.nrcpb@ 123456gmail.com (A.M.S.V.); amolsgene@ 123456gmail.com (A.U.S.); deept_mbi@ 123456yahoo.co.in (D.N.); swatisaxena605@ 123456gmail.com (S.S.); tyagi.anshika9@ 123456gmail.com (A.T.); sureshkumarv1996@ 123456gmail.com (S.V.); nksingh@ 123456nrcpb.org (N.K.S.); trsharma1965@ 123456gmail.com (T.R.S.)
            [2 ]ICAR-Indian Agricultural Research Institute, New Delhi 110012, India; pritam.kalia@ 123456gmail.com
            [3 ]ICAR-Indian Institute of Pulse Research, Kanpur 208204, India; npsingh.iipr@ 123456gmail.com
            [4 ]CCS Haryana Agricultural University, Hisar 125004, India; nryadav@ 123456hau.ernet.in
            Author notes
            [* ]Correspondence: kish2012@ 123456nrcpb.org ; Tel.: +91-011-25841789
            [†]

            Current address: National Agri-Food Biotechnology Institute, Mohali 140306, India.

            Journal
            Genes (Basel)
            Genes (Basel)
            genes
            Genes
            MDPI
            2073-4425
            09 November 2017
            November 2017
            : 8
            : 11
            29120386
            5704226
            10.3390/genes8110313
            genes-08-00313
            © 2017 by the authors.

            Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

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