http://www.100md.com 刘贤华, 白晓东, 仝青英,张韶峰
刘贤华,仝青英, 张韶峰，中国人民武装警察部队总医院科训科中心实验室 北京市 100039
白晓东, 中国人民武装警察部队总医院烧伤整形外科 北京市 100039
刘贤华, 女, 1971-04-19生, 湖北省洪湖市人, 汉族, 1999年重庆市第三军医大学预防医学系本科毕业, 2005年重庆市第三军医大学硕士研究生毕业, 实验师, 主要从事创伤后感染预防和治疗的研究.
国家自然科学基金资助项目, No. 30100197
通讯作者：刘贤华, 100039, 北京市海淀区永定路69号, 北京市武警总医院科训科中心实验室. firstname.lastname@example.org
电话: 010-88276752 传真：010-68211327
Preliminary study on recombination ofJ-HNP-1 and its antibacterial effect in vitro
Xian-Hua Liu, Xiao-Dong Bai,Qing-Yin Tong, Shao-Feng Zhang
Xian-Hua Liu, Qing-YinTong, Shao-Feng Zhang, CentralLabortary, General Hospital of Chinese People’s Armed Police Force,Beijing 100039, China
Xiao-Dong Bai, Departmentof Burn and Plastic Surgery, General Hospital of Chinese People’s ArmedPolice Force, Beijing 100039, China
Supported by NationalNatural Science Foundation of China, No. 30100197
Correspondence to: Dr.Xian-Hua Liu, Central Laboratory, General Hospital of Chinese People’sArmed Police Force, Beijing 100039, China. email@example.com
Received: 2005-07-25Accepted: 2005-08-26
AIM: Toconstruct the recombinant of J chain and human neutrophil peptide-1(HNP-1) gene, and to express it in the mammalian cell system.
METHODS: The J chainand HNP-1 cDNA were ampli-fied from the different plasmids by polymerasechain reaction (PCR), respectively, then the two kinds of cDNA fragmentswere recombined to obtain J-HNP-1 gene by recombinant PCR. After theJ-HNP-1 cDNA fragment was inserted into the mammalian express vectorpcDNA3.1(-)/Myc-HisC, the recombinant vector was transfected to the COS-7cells by lipid transfection. The expression of J-HNP-1 mRNA and proteinwere detected by reverse transcription RCP (RT-PCR) and Western-blot,respectively. The antibacterial activities of cellular soluble protein andculture supernatant were examined in vitro.
RESULTS: The lengths of amplified J chain and HNP-1 were 489 bp and 297 bp, respectively. A band of 786 bp was amplified from COS-7 cells transfected by the recombinant plasmid. Western blot analysis revealed that the lysate of COS-7 cells transfected by rpcDNA3.1(-)/Myc-His /J-HNP-1 had a strong band with molecular weight of about 24 ku. After transfection, the soluble protein and supernatant of the COS-7 cells had significant antibacterial activity, and the diameters of their bacterial inhibition ring were 34 and 43 mm, respectively.
CONCLUSION: The J-HNP-1 recombinant is successf-ully constructed and expressed in vitro, and it has a fa-vorable antibacterial effect.
Key Words:Recombinant PCR; Antimicrobial petide; J chain; Human neutrophil peptide-1
Liu XH, Bai XD, Tong QY, Zhang SF. Preliminary study on recom-bination of J-HNP-1 and its antibacterial effect in vitro. Shijie Huaren Xiaohua Zazhi 2005;13(22):2640-2644
目的:将a-防御素-1(HNP-1)基因与J链基因进行重组使其成为一种新的杀菌肽分子J-HNP-1, 并将其构建于能有效表达和分泌活性J-HNP-1杀菌肽的哺乳动物细胞表达系统 ......