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De-Jellying Embryos

In order to protect the eggs and developing embryos, frogs produce a thick, sticky coating surrounding the egg called the jelly coat. This jelly coat must be removed before the embryo can be effectively manipulated or microinjected.

One thing about completely de-jellied embryos is that they can stick to the bottom of the dish. This can be reduced by adding 0.1% BSA, coating the dish with 1X MBS+0.2%BSA, or raising the embryos in 1% agarose (made in 1/9xMR). To avoid this stickiness problem, you can also raise embryos that are partially de-jellied although these are impossible to micro-inject.

To de-jelly X. tropicalis embryos, immerse in 3% cysteine made up in 1/9xMR (pH to ~7.5-8.0 with NaOH) or water. Swirl the embryos gently and the jelly coat will slowly be removed. NOTE: Embryos being de-jellied before the first cleavage may develop secondary axes if the embryos are swirled too vigorously. Therefore, if the dejellying is being done at the one cell stage, very gentle or no swirling is recommended. For embryos past the first cleavage, de-jelly in an erlenmeyer flask, swirling the embryos every so often to speed the process.

For a partial de-jelly, immerse in cysteine just long enough until the embryos separate and sit next to each other in a monolayer. At this point, the embryos should be easy to manipulate, and the jelly coat should still be visible around the embryo under the stereo-microscope. For a complete de-jelly, immerse in cysteine until the embryos pack tightly with each other without any clear space between the embryos. Again visualization under the stereo-microscope helps.

Once the embryos are adequately de-jellied rinse them 3 - 4 times with 1/9xMR or 0.1X MBS and then sort.

Protocol from the Grainger site

This page contributed by Mustafa K. Khokha, Maura Lane and Timothy C. Grammer