| Summary: | 碩士 === 國立嘉義大學 === 獸醫學系研究所 === 99 === Defensins are one of the antimicrobial peptides. They are thought to act as the first barrier against microorganism invasion and are belonged to innate immunity. They have broad-spectrum antimicrobial activity under a low concentration (20-50 μg/mL) which can destroy bacteria, fungi and some enveloped viruses cell membranes. According to the connectivity of their cysteine residues, defensins are divided into alpha and beta defensin. In healthy animal, alpha-defensin is expressed locally in granulocytes and intestine crypt cells while beta-defensin is expressed in all epitheliums. Synthetic partial cBD-like peptides 1 have been proved to have broad-spectrum antimicrobial activity. Synthetic cBD2 like peptide and cBD3 like peptide have been used to generate antibody against cBD-like peptide 2 and cBD-like ptptied 3 respectively. Studies of cBD1 are limited to mRNA expression level. There is no study about recombinant cBD1 from canine skine. The objective is to express and purify recombinant cBD1 using prokaryotic expression system (E. coli). The purified recombinant cBD1 can be used for cBD1 functional analysis and for generate antibody against cBD1. Recombinant cBD1 is expressed as soluble protien under 0.8 mM IPTG expressed at 16oC for 3 hours. BL21 pET24a-cBD1 has low yield that SDS-PAGE can’t observe the target protein. This system has been proved for protein expression using western blotting. However, how to overcome the low yield of AMPs expression in prokaryotic expression system is still a problem in this study.
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