Induction of asymmetrical cell division to analyze spindle-dependent organelle partitioning using correlative microscopy techniques

Jen Hsuan Wei, Joachim Seemann

Research output: Contribution to journalArticle

10 Scopus citations

Abstract

This protocol describes an assay for the induction of asymmetrical cell division where the entire spindle is segregated into only one of the daughter cells. the procedure consists of four stages: (i) generation of asymmetrical monoasters by arresting cells in early mitosis with a kinesin Eg5 inhibitor; (ii) induction of cell division by microinjection of recombinant Mad1 protein or by the addition of a cdk1 inhibitor; (iii) monitoring the division process by phase-contrast time-lapse microscopy; and (iv) processing for correlative immunofluorescence or correlative electron microscopy. this approach can be applied to determine the requirement for the mitotic spindle in organelle partitioning as well as to investigate the role of the monopolar spindle in cytokinesis. Moreover, the generated nucleus-lacking cytoplast provides an ideal environment to test the feasibility and activity of biological processes in the absence of genomic influence. The protocol takes 2-4 d to complete.

Original languageEnglish (US)
Pages (from-to)1653-1663
Number of pages11
JournalNature Protocols
Volume4
Issue number11
DOIs
StatePublished - Nov 2009

    Fingerprint

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this