Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
???displayArticle.abstract???
Traditionally used for bulk biochemical assays, Xenopus laevis egg extracts have emerged as a powerful imaging-based tool for studying cytoplasmic phenomena, such as cytokinesis, mitotic spindle formation and assembly of the nucleus. Building upon early methods that imaged fixed extracts sampled at sparse time points, recent approaches image live extracts using time-lapse microscopy, revealing more dynamical features with enhanced temporal resolution. These methods usually require sophisticated surface treatments of the imaging vessel. Here we introduce an alternative method for live imaging of egg extracts that require no chemical surface treatment. It is simple to implement and utilizes mass-produced laboratory consumables for imaging. We describe a system that can be used for both wide-field and confocal microscopy. It is designed for imaging extracts in a 2-dimensional (2D) field, but can be easily extended to imaging in 3D. It is well-suited for studying spatial pattern formation within the cytoplasm. With representative data, we demonstrate the typical dynamic organization of microtubules, nuclei and mitochondria in interphase extracts prepared using this method. These image data can provide quantitative information on cytoplasmic dynamics and spatial organization.
???displayArticle.pubmedLink???
34152312
???displayArticle.pmcLink???PMC8752311 ???displayArticle.link???J Vis Exp ???displayArticle.grants???[+]
Belmont,
Real-time visualization of cell cycle-dependent changes in microtubule dynamics in cytoplasmic extracts.
1990, Pubmed,
Xenbase
Belmont,
Real-time visualization of cell cycle-dependent changes in microtubule dynamics in cytoplasmic extracts.
1990,
Pubmed
,
Xenbase
Blow,
Initiation of DNA replication in nuclei and purified DNA by a cell-free extract of Xenopus eggs.
1986,
Pubmed
,
Xenbase
Brownlee,
Importin α Partitioning to the Plasma Membrane Regulates Intracellular Scaling.
2019,
Pubmed
,
Xenbase
Chang,
Robustly Cycling Xenopus laevis Cell-Free Extracts in Teflon Chambers.
2018,
Pubmed
,
Xenbase
Chang,
Mitotic trigger waves and the spatial coordination of the Xenopus cell cycle.
2013,
Pubmed
,
Xenbase
Chatterjee,
In vivo analysis of nuclear protein traffic in mammalian cells.
1997,
Pubmed
Cheng,
Spontaneous emergence of cell-like organization in Xenopus egg extracts.
2019,
Pubmed
,
Xenbase
Deming,
Study of apoptosis in vitro using the Xenopus egg extract reconstitution system.
2006,
Pubmed
,
Xenbase
Desai,
Anaphase A chromosome movement and poleward spindle microtubule flux occur At similar rates in Xenopus extract spindles.
1998,
Pubmed
,
Xenbase
Dunphy,
The Xenopus cdc2 protein is a component of MPF, a cytoplasmic regulator of mitosis.
1988,
Pubmed
,
Xenbase
Field,
Xenopus egg cytoplasm with intact actin.
2014,
Pubmed
,
Xenbase
Field,
Xenopus extract approaches to studying microtubule organization and signaling in cytokinesis.
2017,
Pubmed
,
Xenbase
Field,
Actin behavior in bulk cytoplasm is cell cycle regulated in early vertebrate embryos.
2011,
Pubmed
,
Xenbase
Field,
Assembly of Spindles and Asters in Xenopus Egg Extracts.
2018,
Pubmed
,
Xenbase
Good,
Cytoplasmic volume modulates spindle size during embryogenesis.
2013,
Pubmed
,
Xenbase
Good,
Preparation of Cellular Extracts from Xenopus Eggs and Embryos.
2018,
Pubmed
,
Xenbase
Guan,
Reconstitution of Cell-cycle Oscillations in Microemulsions of Cell-free Xenopus Egg Extracts.
2018,
Pubmed
,
Xenbase
Guan,
A robust and tunable mitotic oscillator in artificial cells.
2018,
Pubmed
,
Xenbase
Hazel,
Changes in cytoplasmic volume are sufficient to drive spindle scaling.
2013,
Pubmed
,
Xenbase
Heald,
Self-organization of microtubules into bipolar spindles around artificial chromosomes in Xenopus egg extracts.
1996,
Pubmed
,
Xenbase
Helmke,
TPX2 levels modulate meiotic spindle size and architecture in Xenopus egg extracts.
2014,
Pubmed
,
Xenbase
Krauss,
Two protein 4.1 domains essential for mitotic spindle and aster microtubule dynamics and organization in vitro.
2004,
Pubmed
,
Xenbase
Lohka,
Formation in vitro of sperm pronuclei and mitotic chromosomes induced by amphibian ooplasmic components.
1983,
Pubmed
,
Xenbase
Minshull,
The A- and B-type cyclin associated cdc2 kinases in Xenopus turn on and off at different times in the cell cycle.
1990,
Pubmed
,
Xenbase
Mitchison,
Roles of polymerization dynamics, opposed motors, and a tensile element in governing the length of Xenopus extract meiotic spindles.
2005,
Pubmed
,
Xenbase
Mitchison,
Bipolarization and poleward flux correlate during Xenopus extract spindle assembly.
2004,
Pubmed
,
Xenbase
Mochida,
Greatwall phosphorylates an inhibitor of protein phosphatase 2A that is essential for mitosis.
2010,
Pubmed
,
Xenbase
Mochida,
Calcineurin is required to release Xenopus egg extracts from meiotic M phase.
2007,
Pubmed
,
Xenbase
Murray,
Cell cycle extracts.
1991,
Pubmed
Murray,
Real time observation of anaphase in vitro.
1996,
Pubmed
,
Xenbase
Murray,
Cyclin synthesis drives the early embryonic cell cycle.
1989,
Pubmed
,
Xenbase
Nguyen,
Spatial organization of cytokinesis signaling reconstituted in a cell-free system.
2014,
Pubmed
,
Xenbase
Nguyen,
Prc1E and Kif4A control microtubule organization within and between large Xenopus egg asters.
2018,
Pubmed
,
Xenbase
Oakey,
Microfluidic Encapsulation of Demembranated Sperm Nuclei in Xenopus Egg Extracts.
2018,
Pubmed
,
Xenbase
Pomerening,
Building a cell cycle oscillator: hysteresis and bistability in the activation of Cdc2.
2003,
Pubmed
,
Xenbase
Sawin,
Mitotic spindle assembly by two different pathways in vitro.
1991,
Pubmed
,
Xenbase
Smythe,
Systems for the study of nuclear assembly, DNA replication, and nuclear breakdown in Xenopus laevis egg extracts.
1991,
Pubmed
,
Xenbase
Wang,
Multiple mechanisms determine ER network morphology during the cell cycle in Xenopus egg extracts.
2013,
Pubmed
,
Xenbase
Wu,
PP1-mediated dephosphorylation of phosphoproteins at mitotic exit is controlled by inhibitor-1 and PP1 phosphorylation.
2009,
Pubmed
,
Xenbase
