Immunoprecipitation of piRNPs and Directional, Next Generation Sequencing of piRNAs

Methods Mol Biol. 2011;725:281-93. doi: 10.1007/978-1-61779-046-1_18.

Abstract

Piwi interacting RNAs (piRNAs) are small (∼25 to ∼30 nucleotide) and are expressed in the germline. piRNAs bind to the Piwi subclade of Argonaute proteins and form the core ribonucleoproteins (RNPs) of piRNPs. We describe a method for the massive identification of piRNAs from immunopurified piRNPs. This strategy may also be used for immunopurification and directional sequencing of RNAs from other RNPs that contain small RNAs.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Drosophila melanogaster / genetics
  • Drosophila melanogaster / metabolism
  • Female
  • High-Throughput Nucleotide Sequencing*
  • Immunoprecipitation*
  • Male
  • Mice
  • Polymerase Chain Reaction
  • RNA, Small Interfering / genetics*
  • RNA, Small Interfering / isolation & purification
  • RNA, Small Interfering / metabolism*
  • Xenopus laevis / genetics
  • Xenopus laevis / metabolism

Substances

  • RNA, Small Interfering