中文摘要：

目的构建苏云金杆菌AiiA基因的真核表达载体并转染A549细胞，观测真核细胞表达的AiiA蛋白对铜绿假单胞菌（Pseudomonas aeruginosa,pa）密度感应（quorum sensing）系统酰基高丝氨酸内酯（N-acylhomoserine lactone，AHL）信号分子及毒力因子合成的影响。方法质粒pET-AiiA经Nhe Ⅰ与Xho Ⅰ酶切后，将AiiA基因片段连接入真核表达质粒pEGFP-N2，脂质体转染A549细胞，提取细胞蛋白，Western blot检测AiiA蛋白表达。将蛋白提取物加入凡培养物中，观察Pa AHL信号分子及毒力因子绿脓素、弹性蛋白酶的生成情况。结果AiiA基因片段成功克隆入真核表达质粒pEGFP-N2，转染该质粒后的A549细胞能够表达AiiA蛋白，且该蛋白能够降低凡培养物中AHL信号分子水平，且绿脓素和弹性蛋白酶的生成减少。结论本研究成功构建了真核表达质粒pEGFP-N2-AiiA，转染细胞后表达的AiiA蛋白能够水解AHL信号分子，减少凡毒力因子的生成，故具有抗Pa感染的潜在价值。

英文摘要：

Objective To construct the eukaryotic expression vector harboring the fragment of AiiA gene, and to investigate the effects of it on the signal of quorum sensing and virulence factors producted by Pseudomonas aeruginosa（Pa）. Methods The plasmid pET-AiiA was cutted by Nhe I and Xho Ⅰ , then the AiiA fragment was cloned into eukaryotic expression vector pEGFP-N2. After the plasmid was transfected into A549 ceils, the protein was extracted and AiiA protein was found in it by Western blot. After the extraction was admixed into the LB broth, from culture supernatant extracts of Pa, the N-acylhomoserine lactone （AHL） was detected by bioassay, and the expression of pyocyanin and elastase were assayed by RT-PCR and optical density. Results The fragment of AiiA gene was cutted and then cloned into pEGFP-N2. AiiA protein was found in the transfected cells. After admixed with the extract harboring AiiA protein, in Pa medium, the AHL was hydrolyzed, and the expression of pyocyanin and elastase were reduced. Conclusion The virulence factors synthesized by Pa were reduced by the AiiA protein expressed in eukaryotic cell.