A FAST AND EFFICIENT MICROINJECTION METHOD IN THE FERTILIZED EGGS OF TELEOST FISH WITH ADHESIVE EGGS

Microinjection is a most popular method in the studies on gene transfer and function in fish. The eggs of teleost are adhesive and once fertilized, the eggs secrete adhesive materials and the eggshell is hardened when contact with water, which makes the microinjection and sampling very difficult. In this study, we developed a fast and efficient microinjection method for the fertilized eggs of teleost fish. The eggshells became non-adhesive when digested in 0.25% trypsin solution (pH 7.1-7.4) at 23℃ for 4min. Furthermore, we compared the new method with the other two - microinjection in the fertilized eggs with intact shells, and in the fertilized eggs with shells completely removed by the trypsin digestion - by injecting Alex-Fluor488-dextran into the fertilized eggs of gibel carp. Compared to the other two methods, the new one generated a number of non-tacky and transparent eggs suitable for injecting and sampling in 5min after fertilization. The new microinjection method established in this study would be a valuable tool for investigating the function of genes involved in the early embryo development and for studying the subtle changes in the nuclear and cytoplasm during the fertilization of teleost fish eggs.