Abstract
This chapter describes the RNA sequencing (RNA-Seq) protocol, whereby RNA from yeast cells is prepared for sequencing on an Illumina Genome Analyzer. The protocol can easily be altered to use RNA from a different organism. This chapter covers RNA extraction, cDNA synthesis, cDNA fragmentation, and Illumina cDNA library generation and contains some brief remarks on bioinformatic analysis.
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References
Nagalakshmi, U., Wang, Z., Waern, K., et al. (2008) The transcriptional landscape of the yeast whole genome defined by RNA sequencing. Science 320, 1344–1349.
Lister, R., O’Malley, R. C., Tonti-Filippini, J., Gregory, B. D., Millar, A. H., and Ecker, J. R. (2008) Highly integrated single-base resolution maps of the epigenome in Arabidopsis. Cell 133, 523–536.
Wilhelm, B. T., Marguerat, S., Watt, S., et al. (2008) Dynamic repertoire of a eukaryotic transcriptome surveyed at single-nucleotide resolution. Nature 453, 1239–1243.
Marioni, J. C., Mason, C. E., Mane, S. M., Stephens, M., and Gilad, Y. (2008) RNA-seq: an assessment of technical reproducibility and comparison with gene expression arrays. Genome Res. 18, 1509–1517.
Parkhomchuk, D., Borodina, T., Amstislavskiy, V., et al. (2009) Transcriptome analysis by strand-specific sequencing of complementary DNA. Nucleic Acids Res. 37, e123.
Wang, Z., Gerstein, M., and Snyder, M. (2009) RNA-Seq: a revolutionary tool for transcriptomics. Nat. Rev. Genet. 10, 57–63.
Mortazavi, A., Williams, B. A., McCue, K., Schaeffer, L., and Wold, B. (2008) Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nat. Methods 5, 621–628.
Shendure, J. (2008) The beginning of the end for microarrays? Nat. Methods 5, 585–587.
Morozova, O., and Marra, M. A. (2008) Applications of next-generation sequencing technologies in functional genomics. Genomics 92, 255–264.
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© 2011 Humana Press
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Waern, K., Nagalakshmi, U., Snyder, M. (2011). RNA Sequencing. In: Castrillo, J., Oliver, S. (eds) Yeast Systems Biology. Methods in Molecular Biology, vol 759. Humana Press. https://doi.org/10.1007/978-1-61779-173-4_8
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DOI: https://doi.org/10.1007/978-1-61779-173-4_8
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Publisher Name: Humana Press
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Online ISBN: 978-1-61779-173-4
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