GenEE project

Professor James S. Harris Group

Solid State Laboratory

Stanford University



GenEE quad-chart




ONR 1aug98 - Brain Imaging (mac ppt 4.0 binhex)
ONR 1aug98 - Brain Imaging (mac ppt 4.0 raw)



GenEE related publications


"The Dynamics of Dentritic Structure in Developing Hippocampal Slices," Michael E. Dailey, and Stephen J. Smith, The Journal of Neuroscience, May 1, 1996, 16(9):2983-2994.


GenEE Images and Movies





Image of descending axon from an Mauthner cell.



Images and Movies

These images illustrate two of the labeling strategies we have developed to allow imaging of the formation of a specific synaptic network connection in the zebrafish.

In movie #1,

Growth Cone,
Growth Cone images

a descending axon from an Mauthner cell neuron is shown growing through an area in which it is known to form synapses onto identified spinal motorneurons.

In movie #2,

Moto Neuron,
Moto Neuron images

one of the target motorneurons is shown as it exhibits the pattern of growth and motility thought to play a role in the process of specific selection of the Mauthner cell partner.

The latest MPEG video sequences ...

below allow a comparison between results obtained the new two-photon system and the older single-photon microscope. The greater clarity and information content of the two-photon images is readily apparent. What is less apparent here but perhaps even more important is the large reduction in physiological photodamage associated with two-photon imaging. With the older single-photon system, any imaging sequence long enough to discern processes of synapse formation inevitably revealed signs of damage and developmental perturbation. With the new two-photon system, such untoward symptoms are rarely or never observed. This instrumental improvement has already allowed observations amounting to a major breakthrough in understanding the forms of active cell motility that initiate specific processes of synapse formation. In addition to pursuing the new experimental opportunities two-photon imaging provides for the substantive study of synaptogenesis, we are working to develop a new spectroscopic detection system which will allow optical signals from multiple distinct fluors (e.g. multiple green fluorescent protein mutants) to be detected simultaneously. Such a detector will greatly advance all of the projects optical imaging components. The two MPEG videos show axons and growth cones from identified brain cells called Mathner neurons growing down the spinal cord of an intact zebrafish embryo. In comparing the quality of images in the two videos, note the greatly increased clarity of the two-photon sequence. The quality of the single photon sequence is limited primarily by the low yield of photons collected during the imaging process. While the low single-photon yields could be increased by increasing ilumination intensity or frame exposure times, such maneuvers inevitably resulted in prompt light-mediated killing of the growth cone. Superior results are obtained by two-photon imaging because of an inherently much more efficient photon collection process and because photoexcitation is always confined to a single, thin optical section during two-photon imaging.

Movie #3 shows


single-photon images of a single axon growing, quick-time version

mpeg version



Movie #4 shows


two-photon images of a single axon growing, quick-time version

mpeg version


The two-photon images show a dramatic increase in resolution, notice the new details!





GenEE Genetic Circuit Diagrams


Genetic circuit diagrams of Cascaded Genetic Links:










Whole/intact fluorescent Drosophila embryo sorting


We have designed and constructed a machine that will automatically separate Drosophila embryos that contain a GFP, (Green Fluorescent Protein), from those that do not. At present, Drosophila is one of the most highly manipulatable genetic model systems; this combined with their fast life cycle results in a high speed at which information can be obtained. Drosophila mutations are maintained over a balancer chromosome and therefore only 25% of the embryos produced from these adults will contain the homozygous mutation. There are a number of Drosophila balancer chromosomes that contain GFP and therefore it is possible to determine visually in living embryos which fraction are the homozygous mutant population. However, in order to obtain some homozygous mutant embryos one would have to hand sort the embryos under a microscope. This is very labor intensive and time consuming. In addition as development occurs very rapidly in Drosophila (within 24 hrs) it would be almost impossible to obtain large enough quantities of sorted embryos at a specific stage in development to do biochemical experiments. We have developed a machine that can separate the GFP containing balancer embryos from the homozygous mutant embryos. This machine can sort the embryos at high speed while maintaining the embryos viability.

For example, a practiced person may be able to isolate 300-400 non- GFP containing embryos in an hour whereas this machine can theoretically isolate over 18,000 an hour. The ability to obtain large amounts of mutant embryos will allow researchers to contain a large number of biochemical and molecular biology studies that have not been possible in the past. It will also allow the analysis of the effects of mutations on the behaviors of fluorescent proteins.


Full-size picture of machine that automatically separates Drosophila embryos




(more to come)