CAGE
Cap Analysis Gene Expression Sequencing
CAGE measures RNA expression and maps TSS in promoters (Takahashi et al., 2012). In this method, RNA is first reverse-transcribed using random primers. The RNA cap and 3′ ends are biotinylated. Nonhybridized, single-stranded RNAs are digested with RNase, leaving 5′ complete cDNAs that are captured using streptavidin beads. The cDNA is processed for sequencing.
Advantages:
- Measures RNA expression levels and maps TSS in promoter regions
- Provides precise mapping of TSS with single-nucleotide resolution
Disadvantages:
- Only works on total mature RNA (Haberle et al., 2016)
- Detection is biased toward TSS of long-lived transcripts (Haberle et al., 2016)
Reagents:
Illumina Library prep and Array Kit Selector
Reviews:
Haberle V. and Lenhard B. Promoter architectures and developmental gene regulation. Semin Cell Dev Biol. 2016;57:11-23
References:
Horie M., Yamaguchi Y., Saito A., et al. Transcriptome analysis of periodontitis-associated fibroblasts by CAGE sequencing identified DLX5 and RUNX2 long variant as novel regulators involved in periodontitis. Sci Rep. 2016;6:33666
Poletti V., Delli Carri A., Malagoli Tagliazucchi G., et al. Genome-Wide Definition of Promoter and Enhancer Usage during Neural Induction of Human Embryonic Stem Cells. PLoS One. 2015;10:e0126590
Andersson R., Gebhard C., Miguel-Escalada I., et al. An atlas of active enhancers across human cell types and tissues. Nature. 2014;507:455-461
Brown J. B., Boley N., Eisman R., et al. Diversity and dynamics of the Drosophila transcriptome. Nature. 2014;512:393-399
Chen Z. X., Sturgill D., Qu J., et al. Comparative validation of the D. melanogaster modENCODE transcriptome annotation. Genome Res. 2014;24:1209-1223
Fort A., Hashimoto K., Yamada D., et al. Deep transcriptome profiling of mammalian stem cells supports a regulatory role for retrotransposons in pluripotency maintenance. Nat Genet. 2014;46:558-566