2.1. Sampling and RNA Isolation

Thirteen healthy Chenfu’s Treefrogs were collected from the Wawu Shan Nature Reserve in Sichuan, China. The experimental frogs were euthanized with 95% alcohol. The liver was dissected and frozen in liquid nitrogen, before freezing at −80 °C for storage until RNA extraction. Trizol^® reagent was used to isolate the total RNA from each sample according to the manufacturer’s protocol (TaKaRa, Otsu, Japan). At the wavelengths of 280 nm and 260 nm, the optical density absorption ratio (Bio-Rad) was measured to determine the RNA concentrations. Specimens with the rate of absorption from 1.7 to 2.2 were used for cDNA synthesis. RNA integrity was measured using gel electrophoresis. Sichuan Agricultural University’s Animal Ethics Committee approved all experiments with live tree frogs with approval number, 71234-2015-0901.