Cellular Localization of Engineered Human LINE-1 RNA and Proteins

Methods Mol Biol. 2016:1400:281-97. doi: 10.1007/978-1-4939-3372-3_18.

Abstract

The human LINE-1 retrotransposon has the ability to mobilize into a new genomic location through an intracellular replication cycle. Immunofluorescence and in situ hybridization experiments have been developed to detect subcellular localization of retrotransposition intermediates (i.e., ORF1p, ORF2p, and L1 mRNA). Currently, these protocols are also used to validate the interaction between retrotransposition complex components and potential cellular partners involved in L1 replication. Here, we describe in details methods for the identification of LINE-1 proteins and/or RNA in cells transfected with vectors expressing engineered human LINE-1 elements.

Keywords: Immunofluorescence; L1 mRNA; LINE-1 retrotransposon; MS2 fluorescent in situ hybridization; ORF1p; ORF2p.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cells, Cultured
  • Fluorescent Antibody Technique
  • Genetic Engineering
  • Humans
  • In Situ Hybridization, Fluorescence
  • Intracellular Space / metabolism
  • Long Interspersed Nucleotide Elements*
  • Open Reading Frames*
  • Protein Biosynthesis*
  • Protein Transport
  • RNA, Messenger / genetics*
  • RNA, Messenger / metabolism*
  • Transcription, Genetic*

Substances

  • RNA, Messenger