Cell-free reconstitution of multi-condensate assemblies

Andrea Putnam, Geraldine Seydoux

Research output: Chapter in Book/Report/Conference proceedingChapter


Biomolecular condensates (BCs) are intracellular condensates that form by phase separation of proteins and RNA from the nucleoplasm or cytoplasm. BCs often form complex assemblies where compositionally distinct condensates wet each other without mixing. In this chapter, we describe methods to reconstitute multi-condensate assemblies from purified components. We include protocols to express, purify, label, and analyze the dynamics of proteins and RNAs that drive multi-condensate assembly. Analysis of the condensation and wetting behaviors of condensates in cell-free reconstituted systems can be used to define the molecular interactions that regulate BCs in cells.

Original languageEnglish (US)
Title of host publicationLiquid-Liquid Phase Coexistence and Membraneless Organelles
EditorsChristine D. Keating
PublisherAcademic Press Inc.
Number of pages31
ISBN (Print)9780128211595
StatePublished - Jan 2021

Publication series

NameMethods in Enzymology
ISSN (Print)0076-6879
ISSN (Electronic)1557-7988


  • FRAP
  • Fluorescence labeling
  • Image analysis
  • Inclusion bodies
  • MBP
  • Maltose binding protein
  • Phase separation
  • Protein purification
  • RNA condensate

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology


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