Literature Search Morpholino oligonucleotides (MOs) have been used as an anti-sense technique to aid researchers in studying development of various vertebrates. In zebrafish, MOs can be used to reduce developmental function, specifically in zygotes, as they can impede splicesomes and ribosome assembly. Additionally they can bind to mRNA to inhibit protein synthesis, thus hindering normal development. This impediments cause blocked gene expression and phenotypic changes in early development. Researches hypothesize that although a good tool, MOs are limited by dilution because of mitosis and protein kinetics. They believe most phenotypic changes in the fish, caused by MOs altering proteins, are displayed within the first three days of development, …show more content…
To begin, MOs were suspended and hydrated in high-grade water, so that they could maintain concentration. Additionally, the solution in which the MOs existed was isotonic, and centrifuged to remove any unwanted particles. The concentration of the MOs was then assessed through spectrophotometry, to allow for proper dosage for injections into the zebrafish embryos. Before injection of the MOs into the embryos, the needle was calibrated with 10 thirty second pulses, and then attached to a foot pedal and placed under a dissecting microscope to allow for equal amounts of injection in each embryo. MOs are injected into the yolk of the embryos to reduce the chances of disruption due to secondary effects in the blastomere stage. The researches injected the embryos between the 1 and 8 cell stage with a dosage of 1.5-6 ng. In this experiment two types of injections were tested, one in which the MOs phenocopied existing mutations already present in the zebrafish, and the other in which MOs were added until a threshold of the reduced wild-type genes was reached. After injection of the MOs the embryos were placed in incubators and incubated at