Large scale chemical mutagenesis screens in zebrafish have led to the isolation of mutations affecting about half the estimated twenty-four hundred genes essential for the development of the fish embryo. The embryonic lethal genes of zebrafish are required for the normal patterning, growth and differentiation of the fish embryo, including its internal organ systems. Some of the genes are required for simple embryonic behaviors. Despite the importance of the chemically induced fish mutants for understanding vertebrate development, a limitation of the chemical screens is that at present it is not possible to clone the mutated genes. This will require the costly development of a zebrafish genome project, followed by positional or candidate-gene cloning approaches. In Drosophilia, a powerful complement to chemical mutagenesis screens has been the use of P-element insertional mutagenesis for rapid cloning of mutant genes. As a complement to chemical mutagenesis in the zebrafish, we have developed a method of insertional mutagenesis for fish using a retroviral vector as the mutagen. In a pilot screen we isolated five embryonic lethal mutations and one adult phenotype and have rapidly cloned three of the six mutated genes. Although less efficient than chemical mutagenesis, the relative ease of cloning mutated genes makes insertional mutagenesis in zebrafish a powerful new technique for performing classical genetics in a vertebrate animal. Hewe we propose to perform a large scale insertional mutagenesis screen in the fish to isolate mutants defective in embryonic development and simple behaviors. We estimate that during the proposed grant period we can isolate approximately four hundred and fifty insertional mutants, roughly fifteen to twenty percent of the embryonic lethals of this organism, and can clone many of the mutated genes. Like the three genes we have cloned to date, many of these will be novel and all are expected to have human homologues. Some of the genes are expected to have important medical implications, including conceivably the diagnosis and treatment of human diseases.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Research Project (R01)
Project #
5R01RR012589-04
Application #
6363983
Study Section
Genome Study Section (GNM)
Program Officer
Chang, Michael
Project Start
1998-02-15
Project End
2003-02-14
Budget Start
2001-06-08
Budget End
2002-02-14
Support Year
4
Fiscal Year
2001
Total Cost
$746,386
Indirect Cost
Name
Massachusetts Institute of Technology
Department
Internal Medicine/Medicine
Type
Schools of Arts and Sciences
DUNS #
City
Cambridge
State
MA
Country
United States
Zip Code
02139
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