RAMPAGE: Promoter Activity Profiling by Paired‐End Sequencing of 5′‐Complete cDNAs
| Autor: | Philippe Batut, Thomas R. Gingeras |
|---|---|
| Rok vydání: | 2013 |
| Předmět: |
Transcriptional Activation
Whole genome sequencing Genetics DNA Complementary High-Throughput Nucleotide Sequencing RNA Promoter General Medicine Biology Article DNA sequencing Gene expression profiling Transcription Initiation Site Promoter Regions Genetic Gene Paired-end tag Illumina dye sequencing |
| Zdroj: | Current Protocols in Molecular Biology |
| ISSN: | 1934-3647 1934-3639 |
| Popis: | This unit introduces RAMPAGE (RNA Annotation and Mapping of Promoters for the Analysis of Gene Expression), a method that harnesses highly specific sequencing of 5′-complete complementary DNAs to identify transcription start sites (TSSs) genome-wide. Although TSS mapping has historically relied on the detection of 5′-complete cDNAs, current genome-wide approaches typically have limited specificity, and provide only scarce information regarding transcript structure. RAMPAGE allows for highly stringent selection of 5′-complete molecules, thus allowing base-resolution TSS identification with a high signal-to-noise ratio. Paired-end sequencing of medium-length cDNAs yields transcript structure information that is essential to interpreting the relationship of TSSs to annotated genes and transcripts. As opposed to standard RNA-seq, RAMPAGE explicitly yields accurate and highly reproducible expression level estimates for individual promoters. Moreover, this approach offers a streamlined 2- to 3-day protocol that is optimized for extensive sample multiplexing, and is therefore adapted for large-scale projects. This method has been applied successfully to human and Drosophila samples, and should in principle be applicable to any eukaryotic system. |
| Databáze: | OpenAIRE |
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