Cloning of Candidate Gene Cdc20 to Characterize the Zebrafish Mutant Zombie
Giacobone, Camille K.
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Zebrafish are an ideal model organism for the study of vertebrate development. The genes responsible for various developmental processes have been identified from a number of genetic screens, such as the Tubingen zebrafish mutagenesis screen in 1996. The specific genetic mutation for an early arrest mutant from the Tubingen screen, zombie, has not yet been identified. The zombie phenotype is lethal and displays abnormal cell nuclei that arrest in mitosis. It has been hypothesized that a mutation in the cell division cycle 20 (Cdc20) gene is responsible for the zombie phenotype. To test Cdc20 as a candidate gene for zombie this study aimed to incorporate a functional copy of Cdc20 into the zombie genome to rescue the zombie phenotype to wild type. To this end, Cdc20 was cloned into a vector commonly used for zebrafish transgenesis. Cloning was performed via in vivo homologous recombination, a method less popular but more efficient than traditional enzymatic based cloning. Although this study was unable to determine if phenotypic rescue was successful, it establishes in vivo homologous recombination as an accessible molecular cloning technique and provides a starting point for further research to test Cdc20 as the gene mutated in zombie. Working to identify the mutation responsible for zombie is important because it has the potential to elucidate more information about the cell cycle and its role in early vertebrate development.