cDNA, genomic sequence cloning, analyzing of ribosomal protein L36A (RPL36A) and its over-expression from giant panda

摘要

The aim of this study is to explore the structural characteristics of the ribosomal protein L36A (RPL36A) gene from giant panda and to investigate its similarities and differences with other species reported. The cDNA and genomic sequence of RPL36A gene were cloned using RT-PCR and touchdown -PCR technology. The results showed that the length of cDNA cloned was 342 bp containing an ORF of 321bp encoded 106 amino acids with an estimated protein molecular weight 12.44kD and a theoretical pI of 11.29. The genomic DNA fragment was 2227 bp containing four exons and three introns. Topology prediction showed that there were one cAMP- and cGMP-dependent protein kinase phosphorylation site, three protein kinase C phosphorylation site, one tyrosine kinase phosphorylation site, one amidation site, one ribosomal protein L44e signature in the RPL36A protein of giant panda. The RPL36A gene could be readily expressed in E. coli. When it was fused with the N-terminally His-tagged protein, it gave rise to accumulation of an expected 17.5kD polypeptide, in good agreement with the predicted molecular weight. The expression product obtained can be purified for studies of its function. Alignment analysis indicated that the nucleotide sequence and the deduced amino acid sequence shared a high homology with other four mammals. In this paper, the cDNA of RPL36A was cloned successfully for the first time from giant panda. The results provided scientific material for enriching and improving the mammals RPL36A gene database.