DeepCAGE Sequencing

A novel approach for expression profiling and promoter identification

The characterisation of the transcriptome comprising tens-of-thousands of genes regulated by various means and differentially expressed at the cellular level is a challenging task for which new approaches are being developed. The first level of transcriptome regulation controls the expression of individual genes, where the amounts of a given transcript can be correlated with the promoter activity of the associated gene.

By incorporating GeneWorks’ Next Generation Sequencing services with DNAFORM’s CAGE (Cap-Analysis-Gene-Expression) technology, GeneWorks provides the powerful new approach of DeepCAGE sequencing for expression profiling and promoter identification.

DeepCAGE sequencing:

• Brings clarity to the heterogeneity and complexity of transcriptomes by changing the view from expression profiling to the "cause of transcription".
• Links expression values to promoter sites for a better understanding on how signaling pathways influence transcription.
• Shifts the focus transcript analysis from 3'-end related information to 5'-ends of mRNA for the identification of Transcriptional Start Sites (TSS).
• Data are unbiased to known transcripts and sequence information.
• Provides experimental validation for transcriptional start sites as computational methods cannot predict end-points of transcripts.

How DeepCAGE sequencing works

CAGE is based on DNAFORM’s Cap-Trapper technology to isolate full-length cDNA. An adaptor is ligated to the 5'-end of full-length cDNAs, which introduces a recognition site for adjacent to the 5'-end of the cDNA is restriction endonuclease that allows for leaving off short tags of 25/27 bp derived from the 5'-end of transcripts.  These short tags are ready for high-throughput sequencing in GeneWorks’ Illumina Genome Analyzer II technology. Several million tags can be sequenced in one lane of the Genome Analyzer. CAGE tags sequenced can then be mapped to genomic sequences, which enable the identification of transcriptional start sites.

For more information about DeepCAGE services please contact GeneWorks at sequencing@geneworks.com.au. Click here for a list of publications relating to use of CAGE technology and see the DNAFORM website for more information about DNAFORM and RIKEN technologies.