Making a New Syngeneic Fish Line Part 1

So for the past 6 months I’ve been following a protocol on how to ‘clone’ or make a syngeneic fish line. This has proved more difficult than anticipated due to the protocol being very deleterious for the embryos (heat shocking them at 46 degrees for 2 minutes!) as well as on the parents….I can now add ‘milking’ fish to my resume! And I cant contact the lab that wrote the protocol as they no longer exist…so any questions I’ve had I’ve had to try and google to no avail and just try and troubleshoot on my own. Needless to say its been beyond frustrating due to all of this. However, over these past 6 months I’ve gotten pretty good at milking the males and females to collect their eggs and sperm without killing them in the process, AND after troubleshooting the protocol and just going with the flow, I’ve successfully done step one of the protocol…which is to make a gynogenetic line of fish, which will then be ‘cloned’ again to make the syngeneic line. These gynogenetic fish are essentially clones of the mother…they have no genetic material from the father. I was super excited when I finally had more than 1 fish live past 14 days…I actually have 12! which isn’t a whole lot, but still pretty happy about it. To give you an idea about why that small number makes me happy, let me tell you that I started out with around 100 good looking eggs…and due to the strenuousness of the protocol, only about 1-5% are supposed to live…well HAH I beat that average! The next step in the process is to let those 12 fish grow to maturity (around 3 months of age) and then use the females to clone again….the babies produced from a single female will be theoretically, syngeneic. This is important to our lab, as we are one of the only Zebrafish labs in the country that already has a fully syngeneic line, which we use to transplant tumor cells into, so if we had more syngeneic lines, we could transplant more tumors and even replace the syngeneic line we already have as the ones I am cloning are completely see-through (they’re albino…the line is called Casper) so they would be preferable for imaging studies, as you would be able to see a tumor growing just by looking at them (well through them really!) and they look much better under microscopes. We are really hoping here in the lab that this works and that we can actually have an optically clear line of syngeneic fish…fingers crossed and wish me luck!

Ashley