Rhodopsin Molecular Evolution in Mammals Inhabiting Low Light Environments

Zhao, Huabin; Ru, Binghua; Teeling, Emma C.; Faulkes, Christopher G.; Zhang, Shuyi; Rossiter, Stephen J.

To verify the coding sequences, we amplified mRNA from the retinal tissue of two non-echolocating fruit bats (Eonycteris spelaea and Rousettus leschenaultii) two high-duty-cycle bats (Rhinolophus ferrumequinum and Hipposideros pratti) and three low-duty-cycle bats (Taphozous melanopogon, Chaerephon plicatus and Myotis ricketti). All of these individuals were collected from China and euthanized as part of a previous project for investigating the animal reservoir of SARS-CoV and in accordance with the guidelines of the China Practice for the Care and Use of Laboratory Animals. Eyes were stored in liquid nitrogen and total RNA isolated using TRIZOL (Invitrogen). First-strand synthesis of cDNA was undertaken using SuperScriptTM II reverse transcriptase (Invitrogen). PCRs mixture included 1 mg of the first-strand cDNA, 0.2 mM of the primers RHFc and RHRc (see Table S2 , Supplementary Material online) and 1 U Taq DNA polymerase (Takara). This yielded a target length of ,1.1 kb. PCR conditions and cloning protocols were the same as those used for genomic DNA.