I. Embryo Preparation
1) Manually dejelly embryo of appropriate stage. Use fine foreps to break hole in hard jelly coat. Remove membrane around embryo by carefully grabbing and pulling with fine forceps. Keep embryos in HBSt containing antibiotics until use.
2) Prepare 1 % agarose-coated operating dish, rinse with HBSt and antibiotics.
3)Transfer donor and recipient embryos into operating dish containing 1X HBSt and antibiotics
4) Dip all tools into 70% EtOH and then into sterile HBSt before using.
5) Use microscalpel or eyebrow knife to cut piece to be grafted fron donor embryos. Remove comparable piece from host embryos. Carefully move graft into position. Use hair loop or forceps to gently push graft into hole in recipient. It may be necessary to push it down several times until it starts to heal. It may be possible to weight the graft down using a small piece of coverslip.
6) After embryos heal from surgery, gently replace 1XHBSt with 50% , then 20 %HBSt solution.