RT Journal Article
SR Electronic
T1 SLIC-CAGE: high-resolution transcription start site mapping using nanogram-levels of total RNA
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 368795
DO 10.1101/368795
A1 Nevena Cvetesic
A1 Harry G. Leitch
A1 Malgorzata Borkowska
A1 Ferenc Müller
A1 Piero Carninci
A1 Petra Hajkova
A1 Boris Lenhard
YR 2018
UL http://biorxiv.org/content/early/2018/07/19/368795.abstract
AB Cap analysis of gene expression (CAGE) is a methodology for genome-wide quantitative mapping of mRNA 5’ends to precisely capture transcription start sites at a single nucleotide resolution. In combination with high-throughput sequencing, CAGE has revolutionized our understanding of rules of transcription initiation, led to discovery of new core promoter sequence features and discovered transcription initiation at enhancers genome-wide. The biggest limitation of CAGE is that even the most recently improved version (nAnT-iCAGE) still requires large amounts of total cellular RNA (5 micrograms), preventing its application to scarce biological samples such as those from early embryonic development or rare cell types. Here, we present SLIC-CAGE, a Super-Low Input Carrier-CAGE approach to capture 5’ends of RNA polymerase II transcripts from as little as 5-10 ng of total RNA. The dramatic increase in sensitivity is achieved by specially designed, selectively degradable carrier RNA. We demonstrate the ability of SLIC-CAGE to generate data for genome-wide promoterome with 1000-fold less material than required by existing CAGE methods by generating a complex, high quality library from mouse embryonic day (E) 11.5 primordial germ cells.