RNA-Seq analysis of the wild barley (H. spontaneum) leaf transcriptome under salt stress

Published Date

May 2015, Vol.338(5):285–297, doi:10.1016/j.crvi.2015.03.010

Molecular biology and genetics/Biologie et génétique moléculaires

Title 

RNA-Seq analysis of the wild barley (H. spontaneum) leaf transcriptome under salt stress

• Author 

• Ahmed Bahieldin ^a,,b,
• Ahmed Atef ^a
• Jamal S.M. Sabir ^a
• Nour O. Gadalla ^a,c
• Sherif Edris ^a,b
• Ahmed M. Alzohairy ^d
• Nezar A. Radhwan ^a
• Mohammed N. Baeshen ^a
• Ahmed M. Ramadan ^a,e
• Hala F. Eissa ^e,f
• Sabah M. Hassan ^a,b
• Nabih A. Baeshen ^a
• Osama Abuzinadah ^a
• Magdy A. Al-Kordy ^a,c
• Fotouh M. El-Domyati ^a,b
• Robert K. Jansen ^a,g

• aDepartment of Biological Sciences, Faculty of Science, King Abdulaziz University (KAU), P.O. Box 80141, 21589 Jeddah, Saudi Arabia
• bDepartment of Genetics, Faculty of Agriculture, Ain Shams University, Cairo, Egypt
• cGenetics and Cytology Department, Genetic Engineering and Biotechnology Division, National Research Center, Dokki, Egypt
• dGenetics Department, Faculty of Agriculture, Zagazig University, 44511 Zagazig, Egypt
• eAgricultural Genetic Engineering Research Institute (AGERI), Agriculture Research Center (ARC), Giza, Egypt
• fFaculty of Biotechnology, Misr University for Science and Technology (MUST), 6th October City, Egypt
• gDepartment of Integrative Biology, University of Texas at Austin, 78712 Austin, USA

Received 19 December 2014. Accepted 12 March 2015. Available online 13 April 2015. 

Abstract

Wild salt-tolerant barley (Hordeum spontaneum) is the ancestor of cultivated barley (Hordeum vulgare or H. vulgare). Although the cultivated barley genome is well studied, little is known about genome structure and function of its wild ancestor. In the present study, RNA-Seq analysis was performed on young leaves of wild barley treated with salt (500 mM NaCl) at four different time intervals. Transcriptome sequencing yielded 103 to 115 million reads for all replicates of each treatment, corresponding to over 10 billion nucleotides per sample. Of the total reads, between 74.8 and 80.3% could be mapped and 77.4 to 81.7% of the transcripts were found in the H. vulgare unigene database (unigene-mapped). The unmapped wild barley reads for all treatments and replicates were assembled de novo and the resulting contigs were used as a new reference genome. This resulted in 94.3 to 95.3% of the unmapped reads mapping to the new reference. The number of differentially expressed transcripts was 9277, 3861 of which were unigene-mapped. The annotated unigene- and de novo-mapped transcripts (5100) were utilized to generate expression clusters across time of salt stress treatment. Two-dimensional hierarchical clustering classified differential expression profiles into nine expression clusters, four of which were selected for further analysis. Differentially expressed transcripts were assigned to the main functional categories. The most important groups were “response to external stimulus” and “electron-carrier activity”. Highly expressed transcripts are involved in several biological processes, including electron transport and exchanger mechanisms, flavonoid biosynthesis, reactive oxygen species (ROS) scavenging, ethylene production, signaling network and protein refolding. The comparisons demonstrated that mRNA-Seq is an efficient method for the analysis of differentially expressed genes and biological processes under salt stress.

Keywords

• Electron transport
• Flavonoid biosynthesis
• Reactive oxygen species

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