DeepCAGE technology and an example of it's usage

DeepCAGE technology and an example of it's usage

I presented the CAGE technology and the paper "Genome-wide detection and analysis of hippocampus core promoters using DeepCAGE" by A.Sandelin et.al. during the Next Generation Sequencing course at Karolinska Insitutet.

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Oxana Sachenkova

April 26, 2013
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  5. RNAseq - High amount of information, not specifically TSS -

    Bias in the outcome: - RNA fragmentation: depleted transcript ends - cDNA fragmentation: biased towards the 3´ends Could other techniques have been used?
  6. ChIPseq Could other techniques have been used? 5´-RACE - H3K4me3

    - RNA Pol II - H3K4me1 - H3K27ac - p300 Active promoters Shen et al, 2012. Nature  Antibody specificity and quality  Lower resolution  Computational peak-calling  ENCODE Consortium provides • ChIP-seq analysis • Dnase hypersensitive site analysis • DNA methylation analysis Used as validation methods for DeepCAGE  Rapid Amplification of cDNA ends (RACE)  Capping of specific transcripts  RT-PCR and PCR needed to amplify the ends of the transcripts (mRNA and cDNA, respectively)  Difficult to validate (designing primers) new core promoters if the exon structure of the downstream protein is not known.  Great sensitivity for lowly expressed transcripts
  7.  Better understanding of the physiologic function of different genes

    in different tissues, or in different cell types within the tissue.  Restore the unfunctional PEP or inhibit the activity of an alternatively activated promoter - antigene RNAs (agRNA) - peptide nucleic acids (agPNA) - locked nucleic acids (LNA) in vitro (Janowski et al. 2005. Nat Chem Biol) General overview of the PEP (Preferentially Expressed Promoters) in a tissue- or cell- specific manner Biological relevance of CAGE  Compare the PEP in normal and disease condition (cancer or other diseases).