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Methods Mol Biol. 2014;1112:33-48. doi: 10.1007/978-1-62703-773-0_3.

High-throughput RNA-seq for allelic or locus-specific expression analysis in Arabidopsis-related species, hybrids, and allotetraploids.

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Section of Molecular Cell and Developmental Biology and Center for Computational Biology and Bioinformatics, The University of Texas at Austin, Austin, TX, USA.


With the next generation sequencing technology, RNA-Seq (RNA sequencing) becomes one of the most powerful tools in quantification of global transcriptomes, discovery of new transcripts and alternative isoforms, as well as detection of single nucleotide polymorphisms (SNPs). RNA-Seq is advantageous over hybridization-based gene quantification methods: (1) it does not require prior information about genomic sequences, (2) it avoids high background problem caused by cross-hybridization, and (3) it is highly sensitive and avoids background and saturation of signals; and finally it is capable of detecting allelic expression differences in hybrids and allopolyploids. We used the RNA-Seq method to determine the genome-wide transcriptome changes in Arabidopsis allotetraploids and their parents, A. thaliana and A. arenosa. The use of this approach allows us to quantify transcriptome from these species and more importantly, to identify allelic or homoeologous-specific gene expression that plays a role in morphological evolution of allopolyploids. The computational pipelines developed are also applicable to the analysis of chromatin immunoprecipitation sequencing (ChIP-seq) data in Arabidopsis-related species, hybrids, and allopolyploids. Comparative analysis of RNA-Seq and ChIP-Seq data will allow us to determine the effects of chromatin modifications on nonadditive gene expression in hybrids and allopolyploids.

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