Posts Tagged ‘microscopy’

Enhancer additivity and non-additivity are determined by enhancer strength in the Drosophila embryo

Jacques Bothma, Hernan Garcia, Samuel Ng, Michael W. Perry, Thomas Gregor, and Michael S. Levine. eLife 2015; 4:e07956 (2015). (more…)

Dynamic Regulation of Eve Stripe 2 Expression Reveals Transcriptional Bursts in Living Drosophila Embryos

Jacques P. Bothma, Hernan G. Garcia, Emilia Esposito, Gavin Schlissel, Thomas Gregor, Michael S. Levine. PNAS 111 (29): 10598–10603 (2014).
(more…)

Quantitative imaging of transcription in living Drosophila embryos links polymerase activity to patterning

Hernan G. Garcia, Mikhail Tikhonov, Albert Lin and Thomas Gregor. Current Biology 23, 2140–2145 (2013).
(more…)

Recent and future projects

Besides these well-advanced projects above there are a number of exciting recently initiated projects:

  1. We started a collaboration with Steve Small (NYU/Biology) to measure the structure-function relationship of one particular enhancer (small piece if DNA that regulates a gene). How is the actual string of code (nucleotides) affecting the final output (mRNA transcripts)? We are genetically engineering enhancer variants that we express in both embryonic cell lines and in living embryos and we are measuring their transcriptional activity by using our single molecule mRNA counting technology.
  2. We are in the process of generating flies that simultaneously express up to three fast-folding/fast-maturing fluorophores attached to different members of the patterning network, and we are building custom microscopes for live imaging that can cope a) with spectral unmixing for multi-color measurements, b) with the fast evolving expression level dynamics, and c) with resolving the full three-dimensional structure of an entire embryo.
  3. In an effort to link our expression level measurements back to evolutionary constraints, we are exploring three interconnected paths: 1) What are the variations of expression levels in natural wild type Drosophila populations from different parts of the world? 2) What are the expression level variations in isogenic fly populations, and how do these variations respond to environmental fluctuations? 3) We are generating tools to quantify phenotypes in the adult organism, such as left-right symmetry of fly wings, to ask how this symmetry is affected if we perturb embryonic expression levels or environmental conditions such as temperature or food supply, and whether perturbances in the embryo and in the adult are correlated.

 

Quantifying the Bicoid morphogen gradient in living fly embryos

Alexander H. Morrison, Martin Scheeler Julien O. Dubuis and Thomas GregorCold Spring Harb Protoc. 2012(4): 398-406 (2012).
(more…)

Researchers Develop Improved Method to Visualize Biologic Molecules

PLoS Press Release for our first paper on mRNA quantification in whole embryos.

“How are biologic molecules arranged inside the embryo so that embryonic development occurs reliably every time? Princeton researchers, led by Thomas Gregor, an assistant professor of physics and the Lewis-Sigler Institute for Integrative Genomics, and Shawn Little, a postdoctoral fellow in the laboratory of Professor Eric Wieschaus in the Department of Molecular Biology, have developed a new method to better understand how an embryo’s basic molecular makeup helps ensure that the embryo’s development occurs reliably every time. The results of this research into the fruit fly Drosophila introduce a method for making precise measurements of biologic units (so-called mRNA molecules) that play a key role in development. The findings are published in the March 1st issue of  in the online, open access journal PLoS Biology.”

(more…)

About the tools we use

For starters, some images:

Laser Scanning Two-Photon Microscope

Laser Scanning Two-Photon Microscope

Detail of a custom built Two-Photon Microscope

Illustration of custom-built two-photon microscope with trans- and epi-detection (also note embryo in special holder).

Illustration of custom-built two-photon microscope with trans- and epi-detection (also note embryo in special holder).