Embryo manipulation/dissection

In this section:

Agarose Plates, 1% (for use with trops or laevis)

Dehydrating embryos

Rehydrating Embryos(for embryos stored in 100% methanol)

Agarose Plates, 1% (for use with trops or laevis)

1 g agarose powder + 100 ml 1/9 X MR without gent in 500 ml jar

make no more than 300 ml (to prevent boil over)

Microwave on high 2 min/100 ml (w/ loose lid)

Swirl and check for solids

When completely melted, fill petri dish 1/4 full

Put in 4C for 5 min, or air dry 30 min

Lift out with tweezers

Can use a mold in agar before it hardens, or cut out desired shape after hardens

Dehydrating embryos

Embryos can be placed directly into 100% methanol in small glass vials.

Embryos can be stored at -20 deg C.

Rehydrating Embryos(for embryos stored in 100% methanol)

Draw off methanol with vacuum

Rehydrate embryos stepwise through each wash, 5 minutes each, on rotating rack:

3:1 methanol:PBS

1:1 methanol:PBS

1:3 methanol:PBS

100% PBS, repeat 2 - 3 times