Clone Of The RPS5 Gene In Yak And Its Prokaryotic Expression

This Gaoyuan Yak was takan as the candidates for the reasch. A pair of primers was designed and synthesized according to the bovine RPS5 gene sequence in Genbank (Accession Number: XM-589792.3). Coding sequence of RPS5 gene was amplified by RT-PCR technology from total RNA of yak's liver, the product was cloned to PMD18-T vector and transfer into competent JM109-cells then got the positive recombinant; tested the sequence and analysed with molecular biology software. Phylogenetic tree of RPS5 protein was constructed. The coding region of Yak's RPS5 gene was cloned to PET28A vector to construct an expression plasmid PET28A/ RPS5. After inducation with IPTG, it groped the best expression conditions of the fusion protein PET28A/ RPS5.The results showed that there was a 1150bp cDNA sequence of RPS5 gene cloned, including 615 bp open reading frame, encoding 204 amino acids. This protein is not secreted protein because we did not find singal peptide via the analysis of amino acid sequences Sequence alignment indicated that there were one mutations in the coding region between yak and bovine, but don't cause amino acid mutation in RPS5 protein. RPS5 protein has high conservation in different species; phylogenetic tree conformed to the laws of species evolution. SDS-PAGE electrophoresis showed that the band with 27 kD of molecular weight was consistent as expected. The result indicated that the successful expression of RPS5 gene。...