蜡样芽孢杆菌总蛋白双向电泳方法的建立和优化

目的探索建立有效的蜡样芽孢杆菌(Bacillus cereus)蛋白质组双向电泳体系,为进一步揭示其促进氧化葡萄糖酸杆菌(Gluconobacter oxydans)产酸的作用机制奠定基础。方法以培养的B.cereus为材料,比较蛋白质制备超声破壁时间、新型细胞裂解液、pH梯度和不同上样量对B.cereus蛋白双向电泳结果的影响。结果与结论采用15 min超声破壁提取B.cereus总蛋白,选用新型蛋白质裂解,用长24 cm、pH 4～7的IPG胶条,采用80μg上样量进行等电聚焦,于60 V 15 min、120 V 6 h条件下进行SDS-PAGE垂直电泳,可以获得背景清晰、重复性好的双向电泳图谱,建立一套用于B.cereus蛋白质组分析的双向电泳方法。     

三种非发酵革兰阴性杆菌的临床分布及耐药性分析

目的:了解临床常见非发酵革兰阴性杆菌的临床分布及耐药性特征,指导临床合理使用抗菌药物,有效控制医院感染.方法:对我院2011-01~12临床分离的非发酵革兰阴性杆菌的临床分布及药敏结果进行回顾性分析.结果:临床分离非发酵革兰阴性杆菌占临床总分离株的20.98%,并且依次以鲍曼不动杆菌、铜绿假单胞菌和嗜麦芽寡养单胞菌为主,对临床常用抗菌药物都存在不同程度耐药性,且耐药形势严峻.结论:重视非发酵革兰阴性杆菌耐药监测,辅助临床制定最佳治疗方案,延缓细菌耐药性发展,控制多重耐药株的流行,避免医院感染暴发流行.     

红霉素3-O-甲基转移酶基因eryG克隆及表达

目的:探索红色糖多孢菌红霉素eryG基因在大肠杆菌中的最佳表达条件.方法:以红色糖多孢菌总DNA为模板,PCR扩增出eryG基因序列,克隆到原核表达载体pET-22b( )上,并使其在大肠杆菌BL21(DE3)中表达.从诱导剂IPTG的终浓度和诱导时间两个因素来优化eryG基因的表达条件.结果:eryG基因可以在大肠杆菌中表达,表达产物EryG经SDS-PAGE分析相对分子质量约为33×103.表达条件经优化后,重组蛋白EryG表达量占菌体总蛋白的55%以上.结论:红色糖多孢菌红霉素eryG基因可以在大肠杆菌中高效地表达,为进一步研究EryG的相关酶学性质、分析其对红霉素发酵产物的影响奠定了基础.     

白介素2工程菌的分批培养及高效表达

观察了培养基成分、pH值、溶解氧以及培养时间和诱导时间对工程菌E.coli K802(pLY-4)生长和rIL-2表达的影响;确定了培养基和发酵控制参数。在此基础上进行了50L发酵罐的分批培养试验,获得了稳定的结果。其中,rIL-2的表达量占菌体总蛋白的50%～70%;每升发酵液rIL-2的产量在300mg以上,经过进一步的纯化和复性,可使rIL-2的纯度达到98%;比活性达到10~7U./mg。     

3种细菌气溶胶特性实验研究

目的了解金黄色葡萄球菌、黏质沙雷菌和枯草芽孢杆菌芽孢的耐冲击能力和耐雾化能力的强弱,观察3种细菌的空气生物学特性。方法使用AGI-30型全玻璃液体冲击采样器(AGI-30)对3种细菌进行耐冲击实验,在12.5 L/min的气流冲击5、10和15 min后测定采样液中细菌滴度和终末采样液体积,采用校正存活率评价3种细菌的耐冲击性。用DV40型气溶胶发生器对3种细菌进行耐雾化实验,分别测定不同雾化时间后的菌液浓度,计算雾化后细菌存活率,评价3种细菌的耐雾化性能。结果采样冲击实验表明,金黄色葡萄球菌的存活率最高,但是在冲击过程中其存活率不稳定,枯草菌芽孢经过15 min的冲击后,存活率为60%,黏质沙雷菌的存活率为72%。在耐雾化能力方面,不同时间的雾化过程中,枯草菌芽孢的存活率最高,经过60 min的雾化后,存活率为83.56%;黏质沙雷菌的相对存活率为40%,金黄色葡萄球菌在雾化过程中存活率状态不稳定。结论金黄色葡萄球菌耐冲击能力最强,但存活率低、稳定性较差;枯草菌芽孢耐雾化能力最强,但耐冲击性能最差,且其特性不适宜作为生物气溶胶的模式菌。3种细菌中,黏质沙雷菌最适于作为生物安全装备生物学评价的指示微生物。     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

目的 对分离自中国东海30 m深海泥的一株产新型大环内酯抗生素(macrolactin s,MLS)的菌株进行形态、生理生化特性及分子水平的分类学研究.方法 通过细菌培养特征观察,菌株生长条件测定、生理生化指标及全细胞脂肪酸成分分析,并特异性扩增其16S rDNA序列,测序比对及构建系统发育进化树,利用本实验室所建立的抗菌模型评价代谢产物的抗菌活性.结果 该菌株与芽孢杆菌属其他种共享97%～99%的保守序列,其中与解淀粉芽孢杆菌(Bacillus amyloliquefaciens)的16SrDNA序列保守度高达99.6%.15:1 iso H/13:0 3OH为该菌株的主要脂肪酸组分.结论 该菌株属于芽孢杆菌属,定名为解淀粉芽孢杆菌MS1721株,其所产大环内酯MLS表现出较强的抗菌活性.     

红色糖多孢菌C-12羟基化酶基因eryK的克隆及原核表达

目的:探讨红色糖多孢菌C-12羟基化酶基因eryK在大肠杆菌中的最佳表达条件.方法:将eryK基因克隆到表达质粒pET-22b( ),使其在大肠杆菌BL21(DE3)中表达,并对装液量、接种量、诱导时间、IPTG浓度和FeCl3浓度等表达条件进行优化.结果与结论:SDS-PAGE电泳显示,在相对分子质量44×103处有EryK蛋白条带,其含量占菌体总蛋白的58%以上.证明高GC含量的红色糖多孢菌C-12羟基化酶基因EryK可以在大肠杆菌BL21(DE3)中高效地表达,为体外研究EryK酶学相关性质以及大规模发酵奠定了基础.     

丙戊酸钠治疗癫痫持续状态的疗效及安全性

目的:评估丙戊酸钠(VPA)快速静滴控制癫痫持续状态(SE)的有效性及安全性.方法:60例SE患者随机分为治疗组和对照组各30例,对照组安定首剂10 mg静脉注射,随后以(1.5～2.0)μg/(kg·min)的速度微泵持续静注安定;治疗组首剂给丙戊酸钠注射剂(10～15)mg/kg,在3～5 min内静脉注射,而后以(0.5～1)mg/(kg·h)持续静脉滴注,根据病情调整滴注速度.结果:治疗组总有效率83.3%,对照组为66.7%,治疗组的有效率高于对照组(P＜0.05).治疗组癫痫完全控制时间(29.01±10.84)s明显短于对照组癫痫完全控制时间[(34.93±10.54)s](P＜0.05).结论: 快速静脉使用VPA治疗癫痫持续状态起效快,作用强,安全有效.     

盐酸二甲双胍壳聚糖-海藻酸钠缓释微囊的制备及性能研究

目的 制备盐酸二甲双胍壳聚糖-海藻酸钠缓释微囊并探索其性能.方法 以壳聚糖和海藻酸钠为囊材,利用复凝聚法将盐酸二甲双胍微囊化.以微囊的药物包封率为制备工艺优化指标,用转篮法测定其体外溶出速率.结果 通过正交实验得出微囊的最佳制备工艺条件为:壳聚糖浓度0.2%,成囊pH 4.5,成囊温度45℃,搅拌速度200r/min.微囊的溶出释放性能实验显示,该载药微囊的累积释放速度远低于市售缓释片剂,在20h左右达到75%左右的最高累积释放值.结论 以最佳制备工艺条件制备含药微囊重现性好、工艺稳定、吸水溶胀性好,体外溶出实验显示具有较好的缓释作用.     

胃CT灌注成像的初步研究

目的:探讨16层CT进行胃CT灌注成像的可行性,评价其在胃肿瘤性病变中的价值.方法:对6例无胃疾病的患者和10例胃肿瘤患者进行了CT灌注扫描.利用胰腺模式进行灌注数据的处理.计算血流量(BF)、血容量(BV)、平均通过时间(MTT)以及表面渗透性(PS).结果:正常胃壁的BF、BV、MTT、PS分别为(13.51±5.34) ml/(min·100g)、(3.31±1.55) ml/100g、(10.40±2.70) s、(3.70±2.86) ml/(min·100g);胃肿瘤的则为(97.32±71.04) ml/(min·100g)、(7.80±2.98) ml/100g、(10.17±3.35) s、(14.80±5.44) ml/(min·100g),P值分别为0.003、＜0.001、0.856和＜0.001.结论:CT灌注成像可提供胃肿瘤的血流动力学信息,在胃肿瘤的诊断中有一定价值.     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

Objective To perform taxonomic studies on a novel macrolactin-producing gram-positive bacterial strain MS1721 isolated from the mud at a depth of 30 m in the East China Sea. Methods Taxonomic studies were conducted based on the phenotypic and BIOLOG analysis, 16S rDNA gene sequencing and blast analysis, and fatty acid components analysis. The antibacterial activity of macrolactin S was determined by evaluation models established in our lab. Results MS1721 strain displayed high similarity with genus Bacillus in similarity values between 97%～99%, in which the nearest neighbor was Bacillus amyloliquefaciens with the similarity value of 99.6%. The predominant fatty acid component was 15:1 iso H/13:0 3OH. Conclusions MS1721 strain belongs to the genus Bacillus, and Bacillus amyloliquefaciens MS1721 was proposed as the name for this strain on the basis of the taxonomic data. The second metabolite-macrolactin s produced by this strain exhibited obvious antibacterial activity.     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

Objective To perform taxonomic studies on a novel macrolactin-producing gram-positive bacterial strain MS1721 isolated from the mud at a depth of 30 m in the East China Sea. Methods Taxonomic studies were conducted based on the phenotypic and BIOLOG analysis, 16S rDNA gene sequencing and blast analysis, and fatty acid components analysis. The antibacterial activity of macrolactin S was determined by evaluation models established in our lab. Results MS1721 strain displayed high similarity with genus Bacillus in similarity values between 97%～99%, in which the nearest neighbor was Bacillus amyloliquefaciens with the similarity value of 99.6%. The predominant fatty acid component was 15:1 iso H/13:0 3OH. Conclusions MS1721 strain belongs to the genus Bacillus, and Bacillus amyloliquefaciens MS1721 was proposed as the name for this strain on the basis of the taxonomic data. The second metabolite-macrolactin s produced by this strain exhibited obvious antibacterial activity.     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

Objective To perform taxonomic studies on a novel macrolactin-producing gram-positive bacterial strain MS1721 isolated from the mud at a depth of 30 m in the East China Sea. Methods Taxonomic studies were conducted based on the phenotypic and BIOLOG analysis, 16S rDNA gene sequencing and blast analysis, and fatty acid components analysis. The antibacterial activity of macrolactin S was determined by evaluation models established in our lab. Results MS1721 strain displayed high similarity with genus Bacillus in similarity values between 97%～99%, in which the nearest neighbor was Bacillus amyloliquefaciens with the similarity value of 99.6%. The predominant fatty acid component was 15:1 iso H/13:0 3OH. Conclusions MS1721 strain belongs to the genus Bacillus, and Bacillus amyloliquefaciens MS1721 was proposed as the name for this strain on the basis of the taxonomic data. The second metabolite-macrolactin s produced by this strain exhibited obvious antibacterial activity.     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

Objective To perform taxonomic studies on a novel macrolactin-producing gram-positive bacterial strain MS1721 isolated from the mud at a depth of 30 m in the East China Sea. Methods Taxonomic studies were conducted based on the phenotypic and BIOLOG analysis, 16S rDNA gene sequencing and blast analysis, and fatty acid components analysis. The antibacterial activity of macrolactin S was determined by evaluation models established in our lab. Results MS1721 strain displayed high similarity with genus Bacillus in similarity values between 97%～99%, in which the nearest neighbor was Bacillus amyloliquefaciens with the similarity value of 99.6%. The predominant fatty acid component was 15:1 iso H/13:0 3OH. Conclusions MS1721 strain belongs to the genus Bacillus, and Bacillus amyloliquefaciens MS1721 was proposed as the name for this strain on the basis of the taxonomic data. The second metabolite-macrolactin s produced by this strain exhibited obvious antibacterial activity.     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

Objective To perform taxonomic studies on a novel macrolactin-producing gram-positive bacterial strain MS1721 isolated from the mud at a depth of 30 m in the East China Sea. Methods Taxonomic studies were conducted based on the phenotypic and BIOLOG analysis, 16S rDNA gene sequencing and blast analysis, and fatty acid components analysis. The antibacterial activity of macrolactin S was determined by evaluation models established in our lab. Results MS1721 strain displayed high similarity with genus Bacillus in similarity values between 97%～99%, in which the nearest neighbor was Bacillus amyloliquefaciens with the similarity value of 99.6%. The predominant fatty acid component was 15:1 iso H/13:0 3OH. Conclusions MS1721 strain belongs to the genus Bacillus, and Bacillus amyloliquefaciens MS1721 was proposed as the name for this strain on the basis of the taxonomic data. The second metabolite-macrolactin s produced by this strain exhibited obvious antibacterial activity.     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

Objective To perform taxonomic studies on a novel macrolactin-producing gram-positive bacterial strain MS1721 isolated from the mud at a depth of 30 m in the East China Sea. Methods Taxonomic studies were conducted based on the phenotypic and BIOLOG analysis, 16S rDNA gene sequencing and blast analysis, and fatty acid components analysis. The antibacterial activity of macrolactin S was determined by evaluation models established in our lab. Results MS1721 strain displayed high similarity with genus Bacillus in similarity values between 97%～99%, in which the nearest neighbor was Bacillus amyloliquefaciens with the similarity value of 99.6%. The predominant fatty acid component was 15:1 iso H/13:0 3OH. Conclusions MS1721 strain belongs to the genus Bacillus, and Bacillus amyloliquefaciens MS1721 was proposed as the name for this strain on the basis of the taxonomic data. The second metabolite-macrolactin s produced by this strain exhibited obvious antibacterial activity.     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

Objective To perform taxonomic studies on a novel macrolactin-producing gram-positive bacterial strain MS1721 isolated from the mud at a depth of 30 m in the East China Sea. Methods Taxonomic studies were conducted based on the phenotypic and BIOLOG analysis, 16S rDNA gene sequencing and blast analysis, and fatty acid components analysis. The antibacterial activity of macrolactin S was determined by evaluation models established in our lab. Results MS1721 strain displayed high similarity with genus Bacillus in similarity values between 97%～99%, in which the nearest neighbor was Bacillus amyloliquefaciens with the similarity value of 99.6%. The predominant fatty acid component was 15:1 iso H/13:0 3OH. Conclusions MS1721 strain belongs to the genus Bacillus, and Bacillus amyloliquefaciens MS1721 was proposed as the name for this strain on the basis of the taxonomic data. The second metabolite-macrolactin s produced by this strain exhibited obvious antibacterial activity.     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

Objective To perform taxonomic studies on a novel macrolactin-producing gram-positive bacterial strain MS1721 isolated from the mud at a depth of 30 m in the East China Sea. Methods Taxonomic studies were conducted based on the phenotypic and BIOLOG analysis, 16S rDNA gene sequencing and blast analysis, and fatty acid components analysis. The antibacterial activity of macrolactin S was determined by evaluation models established in our lab. Results MS1721 strain displayed high similarity with genus Bacillus in similarity values between 97%～99%, in which the nearest neighbor was Bacillus amyloliquefaciens with the similarity value of 99.6%. The predominant fatty acid component was 15:1 iso H/13:0 3OH. Conclusions MS1721 strain belongs to the genus Bacillus, and Bacillus amyloliquefaciens MS1721 was proposed as the name for this strain on the basis of the taxonomic data. The second metabolite-macrolactin s produced by this strain exhibited obvious antibacterial activity.     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

Objective To perform taxonomic studies on a novel macrolactin-producing gram-positive bacterial strain MS1721 isolated from the mud at a depth of 30 m in the East China Sea. Methods Taxonomic studies were conducted based on the phenotypic and BIOLOG analysis, 16S rDNA gene sequencing and blast analysis, and fatty acid components analysis. The antibacterial activity of macrolactin S was determined by evaluation models established in our lab. Results MS1721 strain displayed high similarity with genus Bacillus in similarity values between 97%～99%, in which the nearest neighbor was Bacillus amyloliquefaciens with the similarity value of 99.6%. The predominant fatty acid component was 15:1 iso H/13:0 3OH. Conclusions MS1721 strain belongs to the genus Bacillus, and Bacillus amyloliquefaciens MS1721 was proposed as the name for this strain on the basis of the taxonomic data. The second metabolite-macrolactin s produced by this strain exhibited obvious antibacterial activity.     

一株产新型大环内酯抗生素海洋解淀粉芽孢杆菌的分离鉴定

Objective To perform taxonomic studies on a novel macrolactin-producing gram-positive bacterial strain MS1721 isolated from the mud at a depth of 30 m in the East China Sea. Methods Taxonomic studies were conducted based on the phenotypic and BIOLOG analysis, 16S rDNA gene sequencing and blast analysis, and fatty acid components analysis. The antibacterial activity of macrolactin S was determined by evaluation models established in our lab. Results MS1721 strain displayed high similarity with genus Bacillus in similarity values between 97%～99%, in which the nearest neighbor was Bacillus amyloliquefaciens with the similarity value of 99.6%. The predominant fatty acid component was 15:1 iso H/13:0 3OH. Conclusions MS1721 strain belongs to the genus Bacillus, and Bacillus amyloliquefaciens MS1721 was proposed as the name for this strain on the basis of the taxonomic data. The second metabolite-macrolactin s produced by this strain exhibited obvious antibacterial activity.