CDNA, genomic sequence cloning and overexpression of ribosomal protein S9 gene (RPS9) from Ailuropoda

Ribosomal protein S9 is a component of the 40S ribosomal submit, encoded by the RPS9 gene in eucaryotic cell. It belongs to the S4P family of ribosomal proteins. To explore the structure characteristic of RPS9 gene of giant panda (Ailuropoda melanoleuca), primers were designed according to the known nucleotide sequence of RPS9 genes to clone the cDNA and genomic sequences of RPS9 gene of A. melanoleuca by RT-PCR and PCR strategy respectively, followed by sequencing and analyzing preliminarily. The results indicated that the cDNA fragment of RPS9 from giant panda was 637 bp in size, containing an open reading frame (ORF) of 585 bp, encoding 194 amino acids. The length of the genomic sequence was 6,193 bp, with four exons and three introns. The coding sequence shows a high degree of homology to those of Homo sapiens, Bos taurus, Canis lupus, Mus musculus and Rattus norvegicus by 92.65, 91.11, 92.48, 88.03, and 87.01%, respectively. The homology for the deduced amino acid sequences reached high up to 99.48%. Primary structure analysis revealed that the molecular weight of the putative RPS9 protein was 22.60 kDa, with a theoretical pI of 11.31. Based on topology prediction, there were four distinct types of functional sites in RPS9 protein of giant panda. The RPS9 gene was overexpressed in E. coli, and the N-terminally His-tagged protein was a 28 kDa polypeptide, in good agreement with the predicted molecular weight. The protein obtained could be purified for studying its function further.