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Accession IconERP010515

RNA-seq of Sorghum bicolor with drought tolerant genotype grown under drought-stress or no-stress control environment

Organism Icon Sorghum bicolor
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Technology Badge IconIllumina HiScanSQ

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Description
The present study is expected to reveal differentially expressed genes under drought stress of Sorghum bicolor. The seeds of Sorghum genotype drought tolerant (DT) were grown at 28-32°C day/night temperature with 12/12 h light/dark period in the phytotron glass house. The fully opened uppermost leaves from control and drought stressed seedlings were sampled and stored at -80°C. For RNA-Seq libraries, one microgram of total RNA was extracted with Trizol reagent (Invitrogen, USA) and mRNA libraries were produced using the TruSeq mRNA-Seq library kit (Illumina) according to manufacturer’s instructions. The libraries generated were quantitated using an Agilent Bioanalyzer DNA 1000 chip. (Agilent Technologies, Santa Clara, CA) and a 2x101 cycle paired end sequencing (sequenced by Sandor Pvt. Ltd., Hyderabad, India) was performed using an Illumina HiScanSQ sequencer (Illumina Inc.). Initially, raw reads were processed by NGSQC toolkit (http://59.163.192.90:8080/ngsqctoolkit/) and high quality reads were subjected to de-novo assembly using Trinity assembler (Patel and Jain, 2012). Assembled transcripts were quantified by standard pipeline (Trinity→RSEM→R→DESeq) and those transcripts were removed which has zero FPKM in all four samples (Anders, 2010; Grabherr, et al., 2011; Li and Dewey, 2011). These transcripts were further processed by transdecoder tool to retrieve full length coding sequence and subsequent annotated by FastAnnotator (http://fastannotator.cgu.edu.tw/index.php) (Chen, et al., 2012). Pathway enrichment analysis was performed for the predicted transcripts by KEGG Automatic Annotation Server (KAAS; www.genome.jp/tools/kaas/) for the classification of spatial and temporally governed pathways.
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