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. 2021:2296:263-278.
doi: 10.1007/978-1-0716-1358-0_16.

Coupled Transcriptomics for Differential Expression Analysis and Determination of Transcription Start Sites: Design and Bioinformatics

Antonio Rodríguez-García  1 , Alberto Sola-Landa  2 , Rosario Pérez-Redondo  2
Affiliations

Coupled Transcriptomics for Differential Expression Analysis and Determination of Transcription Start Sites: Design and Bioinformatics

Antonio Rodríguez-García et al. Methods Mol Biol. 2021.

Abstract

The term coupled transcriptomics is coined to describe a design of an RNA-seq experiment intended for both differential expression analysis and genome-wide determination of the transcription start sites (TSS). The minimal requirements for the first analysis are two experimental conditions with at least two biological replicates enabling statistical tests. The second analysis involves the bioinformatics comparison of the data generated from a control RNA-seq library with another library enriched in primary transcripts using Terminator™ 5'-phosphate-dependent exonuclease, in an experiment denominated differential RNA-seq (dRNA-seq). Usually, dRNA-seq is carried out with specific protocols for library construction, different of those used for common differential expression analysis. Our experimental design allows to use the same data for both analyses, reducing the number of libraries to be generated and sequenced. This is a guide for designing a coupled transcriptomics experiment and for the subsequent bioinformatics procedures. The proposed methods can be applied to the detection and study of small RNA genes.

Keywords: Differential expression; RNA-seq; TSSpredator; Transcription start site; Transcriptomics; dRNA-seq; sRNA.

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References

    1. Bainbridge MN, Warren RL, Hirst M et al (2006) Analysis of the prostate cancer cell line LNCaP transcriptome using a sequencing-by-synthesis approach. BMC Genomics 7:246. https://doi.org/10.1186/1471-2164-7-246 - DOI - PubMed - PMC
    1. Emrich SJ, Barbazuk WB, Li L et al (2007) Gene discovery and annotation using LCM-454 transcriptome sequencing. Genome Res 17:69–73. https://doi.org/10.1101/gr.5145806 - DOI - PubMed - PMC
    1. Lister R, O’Malley RC, Tonti-Filippini J et al (2008) Highly integrated single-base resolution maps of the epigenome in arabidopsis. Cell 133:523–536. https://doi.org/10.1016/j.cell.2008.03.029 - DOI - PubMed - PMC
    1. Nagalakshmi U, Wang Z, Waern K et al (2008) The transcriptional landscape of the yeast genome defined by RNA sequencing. Science 320:1344–1349. https://doi.org/10.1126/science.1158441 - DOI - PubMed - PMC
    1. Croucher NJ, Thomson NR (2010) Studying bacterial transcriptomes using RNA-seq. Curr Opin Microbiol 13:619–624 - DOI

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