Three transcriptomes from Tibetan Loaches reveals accelerated evolution and adaptation to the Tibetan Plateau
An adult male Triplophysa scleroptera was collected in upper reaches of the Yellow River in Ruoergai County of Sichuan Province. The fish was netted directly into an ice water bath until no movement was dissected (<30 seconds). Total RNA from five tissues (heart, liver, brain, spleen and kidney) was extracted using SV Total RNA Isolation System (Premega). Equal quantities of RNA from each tissue were pooled to generate one mixed sample. Then, the mRNA was fragmented using divalent cations under elevated temperature in NEBNext First Strand Synthesis Reaction Buffer (5X). First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase (RNase H-). Second strand cDNA was subsequently synthesized. Sequencing library was generated using TruSeq SR Cluster Kit v3-cBot-HS (Illumina) following manufacturer’s recommendations. In the final step, the library preparations were sequenced on an Illumina Hiseq 2000 platform and 100 bp paired-end reads were generated.