重组大肠杆菌利用乳糖自诱导系统融合表达天蚕素AD和蛙BuforinⅡ

以包含质粒pET-Trx-CAD-BuforinⅡ的大肠杆菌BL21(DE3)为研究对象,开发了一种利用乳糖自诱导方式表达天蚕素AD和蛙BuforinⅡ的表达系统。首先,在LB培养基中研究了乳糖浓度、诱导时机和诱导时间对目的蛋白表达的影响。然后,利用"分解代谢物阻遏"原理构建乳糖自诱导表达系统,以"葡萄糖+主碳源"两种碳源的依次代谢将蛋白表达过程分为两个阶段:第一阶段为菌体生长阶段,菌体利用葡萄糖生长,乳糖无法进入细胞诱导蛋白表达;第二阶段为蛋白表达阶段,葡萄糖耗尽,菌体开始利用主碳源生长;同时乳糖进入细胞诱导蛋白表达。最后,利用优选的"葡萄糖+木糖"组合自诱导表达天蚕素AD和BuforinⅡ的融合蛋白,得到的目的蛋白占全菌总蛋白含量的45.2%。     

科技新闻媒体关注指数排行榜

(新闻时段:2011-06-01 至 2011-06-10;为新闻关注度,☆为/2)1建立肠出血性大肠杆菌检测法 [关注指数:]7 日,中国疾病预防控制中心传染病预防控制所、南开大学泰达生物技术学院合作,     

为什么生命是有限的？

生物为什么会衰老和死亡？人终有一死,没什么疑义。小至老鼠,大至巨象,终将迎来生命终结的时刻。不过,对草履虫、大肠杆菌等通过分裂完成增殖的生物来说,有“死亡”的时刻吗？它们的生命通过分裂无限延续,看起来永远不会死,但事实并非如     

A novel gene <Emphasis Type=Italic>R049</Emphasis> identified in uropathogenic <Emphasis Type=Italic>Escherichia coli</Emphasis> provides partial protection in mice from colonization

Uropathogenic Escherichia coli (UPEC) is the most common causative organism of human urinary tract infection (UTI). Several UPEC virulence factors have been identified, but more are yet to be found. We previously identified a novel 789-bp-long DNA fragment (named R049) in UPEC strain 132 using a suppressive subtractive hybridization technique. In the present study, we used genome walking to elongate the sequence of this fragment to obtain the whole gene sequence and examined the role of this gene product in generating protective immunity. Through bioinformatic analysis, we predicted that this gene is a 1311-bp open reading frame (ORF), which we designated ORF_R049 (GenBank accession No.: EF488001). We further constructed a prokaryotic expression system to express full recombinant R049 protein and isolated and purified the protein through IPTG induction and nickel affinity chromatography. Using mouse immunosera generated by the purified protein, we confirmed the natural expression and outer membrane localization of the protein in wild-type strain UPEC132 by Western blotting. To test the potential of this protein as a vaccine candidate, we immunized mice with the recombinant protein before challenging them with UPEC132 through the urinary tract. The results showed significantly reduced bacterial colonization in the urine and kidneys of the immunization group compared with the control group. However, the degree of renal pathological damage was not significantly improved in the immunized mice. Our study has identified a novel gene of UPEC which can generate protective immunity against UTI. This novel gene provides a promising new vaccine candidate.     

论大肠杆菌的防治

大肠杆菌是动物肠道正常菌系的主要成员,为革兰氏阴性、中等大小的杆菌,无明显的夹膜,不形成芽孢,有鞭毛能运动,变异菌株无鞭毛不能运动。根据致病力特点分为:致病性菌株、条件致病性菌株和非条件致病性菌株。根据表面抗原O、K和鞭毛抗原H以及菌毛抗原F,大肠杆菌形成了复杂的血清型,在防治方面困难很大。大肠杆菌对热的抵抗力不强,对消毒药敏感,常规浓度短时间即可杀灭。     

响应面法优化大肠杆菌表达Hyperthermus butylicus耐热醛缩酶诱导条件的研究

为了提高重组耐热2-脱氧核糖-5-磷酸醛缩酶的表达水平,该文基于中心组合实验的响应面分析方法,对基因工程E.coli的诱导条件进行了优化,具体考察因素包括诱导剂(异丙基-β-D-硫代半乳糖,IPTG)浓度和诱导时机.实验结果表明该方法可以提高目标蛋白的表达量,依据回归分析确定了最佳诱导条件为:IPTG浓度0.57 mM...     

北豆根生物碱抑菌作用的微量量热法研究

从中药北豆根中提取北豆根生物碱.应用微量量热仪分别测定了不同浓度的北豆根生物碱对大肠杆菌、金黄色葡萄球菌、枯草杆菌代谢作用的热功率—时间曲线,运用Logistic方程计算出细菌的生长速率常数,建立了生长速率常数与药物浓度间的关系,进而确定了最佳抑菌浓度.北豆根生物碱对大肠杆菌、金黄色葡萄球菌、枯草杆菌的最佳抑菌浓度分别为0.7977mg/mL、0.2089mg/mL、0.1129mg/mL.通过对三个最佳抑菌浓度数据的比较,可知北豆根生物碱对三种细菌抑制效果为:枯草杆菌金黄色葡萄球菌大肠杆菌.     

TiO2-HPA薄膜抑制大肠杆菌作用研究

用溶胶-凝胶法制备了不同浓度、不同层数的掺有杂多酸的TiO2薄膜,研究了HPA浓度、镀膜层数对薄膜抑菌性能的影响。结果表明：掺杂1.25%HPA的TiO2-HPA薄膜的抑菌效果较好,随着镀膜层数的增加,膜的抑菌性能逐步提高。