Fluorescence-Activated Nucleolus Sorting in Arabidopsis

Methods Mol Biol. 2016:1455:203-11. doi: 10.1007/978-1-4939-3792-9_15.

Abstract

Nucleolar isolation allows exhaustive characterization of the nucleolar content. Centrifugation-based protocols are not adapted to isolation of nucleoli directly from a plant tissue because of copurification of cellular debris. We describe here a method that allows the purification of nucleoli using fluorescent-activated cell sorting from Arabidopsis thaliana leaves. This approach requires the expression of a specific nucleolar protein such as fibrillarin fused to green fluorescent protein in planta.

Keywords: Arabidopsis thaliana; FACS; Fibrillarin; Nucleolus.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Arabidopsis / metabolism*
  • Cell Nucleolus / metabolism*
  • Chloroplasts / metabolism
  • Chromosomal Proteins, Non-Histone / genetics
  • Chromosomal Proteins, Non-Histone / metabolism
  • Flow Cytometry* / methods
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Microscopy, Fluorescence
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism

Substances

  • Chromosomal Proteins, Non-Histone
  • Recombinant Fusion Proteins
  • fibrillarin
  • Green Fluorescent Proteins