Using RAMPAGE to Identify and Annotate Promoters in Insect Genomes

: Application of Transcription Start Site (TSS) profiling technologies, coupled with large-scale next-generation sequencing (NGS) has yielded valuable insights into the location, structure, and activity of promoters across diverse metazoan model systems. In insects, TSS profiling has been used to characterize the promoter architecture of Drosophila melanogaster (Hoskins et al., Genome Res 21(2):182-192, 2011) and subsequently was employed to reveal widespread transposon-driven alternative promoter usage in the fruit fly (Batut et al., Genome Res 23:169-180, 2012).In this chapter we discuss the computational analysis of the experimental data derived from one of TSS profiling methods, RAMPAGE (RNA Annotation and Mapping of Promoters for Analysis of Gene Expression) that can be used for the precise, quantitative identification of promoters in insect genomes. We demonstrate this using the software tools GoRAMPAGE (Brendel and Raborn, GoRAMPAGE-A workflow for promoter detection by 5'-read mapping. https://github.com/BrendelGroup/GoRAMPAGE , 2016) and TSRchitect (Raborn and Brendel, TSRchitect: promoter identification from large-scale TSS profiling data. R Bioconductor package version 1.8.0 [Online]. Available: http://bioconductor.org/packages/release/bioc/html/TSRchitect.html , 2017), providing detailed instructions with the aim of taking the user from raw reads to processed results.