Zebrafish functional genomics development at UW-Stout

Abstract

Since the completion of the human genome sequencing project, morpholino phosphorodiamidate oligonucleotide (MO) knockdown in zebrafish has been increasingly used to elucidate human gene function. As part of the effort to expand the functional genomics screening capacity at the University of Wisconsin-Stout, this project investigated MO microinjection techniques, embryo production, and brine shrimp survival. One-to two-cell embryos injected with the pigment-inhibiting tyrosinase MO were observed at 48 hours post fertilization for pigmentation. Injection efficiency was calculated by dividing the number of zebrafish lacking pigment by the total number of injected zebrafish. To obtain sufficient embryos for MO experiments, the effects of a dry food diet and live (brine shrimp) food diet on embryo production were analyzed. Effects of a yeast diet on brine shrimp survival were also studied to provide zebrafish with healthy brine shrimp. Results included a 92% injection efficiency, greater embryo production with the dry food diet, and increased brine shrimp survival with a yeast diet. This work provided an important foundation in the development of a reverse-genetic screen for future students.