牛气管抗菌肽在山羊乳腺组织中的表达

Expression of Bovine Tracheal Antibacterial Peptide in Mammary Gland of Goats

根据已经发表的牛气管抗菌肽(bTAP)cDNA序列设计4条短的寡核苷酸片段,以重叠延伸PCR的方法合成bTAP基因,将其克隆至pMD-18T载体,序列测定正确后,构建bTAP基因的乳腺组织特异性表达载体pBcp-bTAP,将其溶于生理盐水后直接注射山羊乳腺小叶,使bTAP基因在乳腺组织内表达,收集注射质粒前后的奶样,测试奶样对金黄色葡萄球菌,大肠杆菌的抑制能力,结果显示注射pBcp-bTAP后的奶样表现出对金黄色葡萄球菌和大肠杆菌具有明显的抑制作用,这种抑制活性可以持续至20h以上。

According to the sequence of bovine tracheal antibacterial peptide, a codon-optimized gene was synthesized by overlap extension PCR. The gene was then cloned into the vector pMD-18T, the recombinant clones containing the gene were confirmed by DNA sequencing. In order to express bTAP in animal mammary gland, the mammary specific expression vector containing the goat β-casein gene 5＇ flanking regulation sequence, the gene of bTAP and the sequence of bovine growth hormone polyadenylation signal was constnacted. The vector was injected directly into the lactating mammary gland of goats. The expression and bactericidal activity were detected using inhibition zone assay for two bacteria： E.coli K12D31 and Staphylococcus aureus. Bovine tracheal antibacterial peptide expressed in goat milk possesses bactericidal activity against these two kinds of bacteria.