Zebrafish have many characteristics that make it an ideal model system, said Collodi. They have a large number of embryos available — 2,000 in a year compared to just a few dozen from a mouse. This is advantageous because it both speeds up research progress and gives a larger sample population to work with. Zebrafish eggs are transparent, which allows researchers to see every cell develop from fertilization onward. In mice, the embryo is in the mother, which prevents observation at every stage of development.
In genetic research, "knockouts" are used. These are varieties of organisms that have specific mutations introduced to them. The organisms are then studied to see how the genetic change affects development.
"The only downside to working with zebrafish, thus far, has been the inability to introduce target mutations for genetic research and have those mutations carry over to future generations," he said. "This has been a problem with every organism except mice, which can carry mutations through the germ line."
The problem preventing success, with creating knockouts, lies in the characteristics of embryonic stem cells. Stem cells are unspecified cells that develop into different types of cells and tissues. Once a mutation is introduced into the genome, stem cells must still be able to properly differentiate. Collodi's team is developing a way to grow stem cells that stay pluripotent, or unspecialized, longer.
"We are optimizing the model system so we can maintain a larger percent of the culture at the pluripotent level," said Jennifer Crodian, research associate. "This will allow us more time to do genetic manipulations."
Right now, the group has one type of knockout raised to almost three months old, which is nearly mature enough to screen to see if the mutation is, in fact, in the germ line. If they are successful, other knockouts will be engineered depending on the interests of researchers.
For example, Collodi said his team would particularly like to knock out the genes that code for a class of proteins responsible for controlling the movement and differentiation of cells in the embryo. This would allow the researchers to further refine their methods.
Pending unforeseen problems, the research team could see success quite soon. This is beneficial to people because while zebrafish won't replace the use of mice, it will provide an additional, easier assay for studying human disease, said Collodi.
The National Institutes of Health grant helped tremendously with the progress of the research, said research associate Lianchun Fan, who is most responsible for the project and obtaining the grant. It has allowed them to use better equipment and continue on in a timely fashion.
