Molecular cloning of human IL-16 gene and its expression in Escherichia Coli (34.17)

Abstract

Abstract Objective: To clone human IL-16 gene, and construct its prokaryotic expression vector, and express it in E. coli DH5. Methods: The total RNA was extracted from peripheral blood mononuclear cells stimulated by histamine. And hIL-16 cDNA was amplified by using RT-PCR and cloned into pUC18 T-vector. After the sequence of hIL-16 cDNA was confirmed, the cDNA was inserted into prokaryotic expression vector pMAL-C2. The recombinant expression plasmid pMAL-IL-16 was identified by endonucleases digestion and PCR, and then transformed into E. coli DH5, human IL-16 expressed in DH5 was identified by SDS-PAGE and Western blotting method. Results: Obtained cDNA of hIL-16 was identical with that published on Genebank, and the prokaryotic expression vector pMAL-IL-16 was constructed correctly. Recombinant protein was expressed in E.coli when induced with IPTG and had a molecular weight of 60kD, wich is consistent with theoretical calculation. Conclusion: The hIL-16 cDNA was cloned and the prokaryotic expression plasmid pMAL-IL-16 was constructed successfully. The E. coli DH5 that stably expressed hIL-16 was obtained, which provide experimental basis for the further studying of functional activities and application of hIL-16. [Key words] Human interleukin-16, Molecular cloning, Prokaryotic expression