MicroRNA cloning from cells of the immune system

Methods Mol Biol. 2010:667:67-77. doi: 10.1007/978-1-60761-811-9_5.

Abstract

MicroRNAs have emerged as - important posttranscriptional regulators of gene expression. Small RNA cloning is a powerful method to identify new microRNAs (miRNAs) and to profile miRNA expression. In addition, it reveals end heterogeneity that may be important in miRNA function. Here, we describe a protocol that is optimized to clone small RNAs from limited amounts of starting material. This is often the case for studying miRNAs in a highly purified population of immune cells or other primary cell types with limited numbers. The small RNAs cloned with this protocol will have a 5'-PO(4) and 3'-OH group, typical features of miRNAs, so majority of the cloned small RNAs will be miRNAs.

MeSH terms

  • Animals
  • Cloning, Molecular / methods*
  • Humans
  • Immune System / anatomy & histology*
  • Immune System / physiology*
  • MicroRNAs / genetics*
  • Nucleic Acid Amplification Techniques / instrumentation
  • Nucleic Acid Amplification Techniques / methods
  • Nucleic Acid Hybridization / methods
  • Polymerase Chain Reaction / instrumentation
  • Polymerase Chain Reaction / methods

Substances

  • MicroRNAs