Time Lapse Imaging of Zebra fish Embryonic Development and
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Transcript Time Lapse Imaging of Zebra fish Embryonic Development and
Cell Division Dynamics
A Closer Look at Gastrulation in Zebrafish
Salome Chuang, Daniel Holland-Moritz
Mentor: Luca Caneparo
Summer Research Connection
Oak Crest Institute of Science
California Institute of Technology
July 30, 2010
Cell Division: A Crucial Biological
Process
• Normal:
• Growth
• Development
• Tissue repair
• Abnormal:
• Cancer
http://tainano.com/chin/Molecular%20Biology%20Glossary.htm
An Introduction to Cell Division
Our Project: Dynamics of Cell
Division in Embryos
End of Cell Division
Beginning of Cell Division
Spindle Poles
Difficulties of Studying Cell Division
in Live Embryos
• Ethical concerns (humans)
• Opaque embryos
• Development Rate
• Production Rate
• Manipulation
• Storage/Care
• External vs. Internal
Development
Images from Kidscientist.com and odec.ca
Zebrafish: An Ideal Model for
Studying Embryonic Cell Division
Very common for use in research because of:
Rapid reproductive rate
Transparent embryos
Rapid development
Easily cared for and contained
Easy to induce and screen for mutations
Genome almost fully sequenced
Bar: 250 µm
Images from focusonnature.be
A Brief History on Zebrafish
George Streisinger
Established zebrafish as an ideal subject for imaging and
manipulation.
Very small species of minnow
Native to the Ganges region of India
Named for distinctive stripes
Since then zebrafish have been popularized as a model for
studying vertebrate development:
Cell biology and development
Organ system development
Genetic studies
Image from www.zfin.org
Stages of Zebrafish Development
Key:
=Injection stage
=Imaging stage
Shield Stage to Gastrulation
Cell Division in Dorsal Region of
Embryos
Dorsal
Posterior
Significance of the Dorsal Region
Dorsal region becomes...
Nervous system
Application:
Cell orientation could have an impact on brain and
nervous system development at an early age
Cell imaging provides better insight into....
Embryonic development
Cell division mechanisms and signaling in vertebrates
Shield Stage to Gastrulation: Trends
Tissue differentiation
Dorsal region: Definite pattern
Cells divide along the anterior-posterior plane
Concha, Adams, 1998
Visualizing Cell Division Dynamics
End of Cell Division
Beginning of Cell Division
Spindle Poles
Labeling of Cell Structures to Study
Cell Division Dynamic
Red: Nucleus
Green: Cytoskeleton
Methods
Genetic Engineering of Fluorescent Embryos
mRNA codes for fluorescent
protein in cytoskeleton and
nucleus
DNA RNA Protein
Synthetic mRNA produced
for injection
mRNA injected into embryos
at one cell stage
Image from protein-dynamics.l001.de
Methods
Imaging of Embryos Using Confocal Microscopy
• Confocal microscope detects fluorescence
• Embryos oriented to image dorsal region
• Anterior and posterior marked
• Imaged over a period of 4 hours
3D Image Reconstruction
Viewing Cell Divisions in Live
Embryos
Close Up: Cell Division
Viewing Cell Divisions with Spindle
Fibers Labeled
Tracking Cell Rotation During
Division: Before and After
3D Image Navigation
Analysis
Special computer program tracks division alignment
Point assigned to each half of
separating chromosomes
Program tracks and measures the
angle of division relative to the
anterior-posterior plane and dorsalventral plane
Image Courtesy of L. Caneparo
Results
We observed an active rotation of the cells to align
prior to division.
Before
After
Future Directions
• Further Analysis
• Quantify angle/character of rotation
• Statistically analyze data
• Investigate cell signaling patterns
• Compare cell division in other vertebrate models
• Explore implications on neural development
Acknowledgements
Special Thanks to:
• Luca Caneparo
• Fraser Lab
• Sherry Tsai
• James Maloney
• California Institute of Technology
• Howard Hughes Medical Institute
• Siemens Foundation
• Our families