Detecting expressed genes using CAGE

Methods Mol Biol. 2014;1164:67-85. doi: 10.1007/978-1-4939-0805-9_7.

Abstract

Cap analysis of gene expression (CAGE) provides accurate high-throughput measurement of RNA expression. By the large-scale analysis of 5' end of transcripts using CAGE method, it enables not only determination of the transcription start site but also prediction of promoter region. Here we provide a protocol for the construction of no-amplification non-tagging CAGE libraries for Illumina next-generation sequencers (nAnT-iCAGE). We have excluded the commonly used PCR amplification and cleavage of restriction enzyme to eliminate any potential biases. As a result, we achieved less biased simple preparation process.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkaline Phosphatase / metabolism
  • Animals
  • Base Sequence
  • Biotinylation / methods
  • DNA, Complementary / genetics*
  • DNA, Complementary / isolation & purification
  • DNA, Complementary / metabolism
  • Exodeoxyribonucleases / metabolism
  • Gene Expression Profiling / methods*
  • Gene Library
  • High-Throughput Nucleotide Sequencing / methods*
  • Humans
  • Mice
  • Promoter Regions, Genetic
  • RNA / genetics*
  • RNA / metabolism
  • Reverse Transcription
  • Ribonuclease, Pancreatic / metabolism
  • Transcription Initiation Site*

Substances

  • DNA, Complementary
  • RNA
  • Exodeoxyribonucleases
  • exodeoxyribonuclease I
  • Ribonuclease, Pancreatic
  • Alkaline Phosphatase