For DNA extraction the animal was thawed, weighed and Prepman Ultra (Applied Biosystems) was added directly to the tube at the following rates: #0.20 g, 450-500 mL; 0.21-0.30 g, 500-750 ml; .0.31 g, 900 mL. This was followed by approximately 100 mL sterile 1.0 mm zirconia silicone beads (BioSpec Products). The tadpole was homogenised in a mini-beadbeater (BioSpec Products) for 30 sec, microfuged at 13,0006 g for 30 sec then homogenised in a mini-beadbeater for a further 30 sec. To extract DNA the sample was heated to 100uC for 20 min, left to stand for 4 min at room temperature (RT), microfuged at 13,0006 g for 6 min and the DNA (350-650 ml) transferred to a clean tube.