大肠埃希菌氨基糖苷类耐药株 Aac(3) Ⅱ 基因保守区分析

常规方法分离鉴定47株大肠埃希菌,以标准纸片扩散法(K B法)对其进行常用氨基糖苷类药物敏感性测定,耐药株经PCR检测aac(3) Ⅱ基因保守区,扩增产物进行DNA测序分析。初步探讨大肠埃希菌氨基糖苷类抗生素耐药株与aac(3) Ⅱ基因保守区之间的关系,结果显示本地区大肠埃希菌氨基糖苷类抗生素耐药株的aac(3) Ⅱ基因保守区具65位G、84位T和65位A、84位C两种基因型,且高度耐药菌株皆为65位G、84位T基因型。Abstract:According to standard K B method,bacteriostatic tests were performed to screen out aminoglycoside resistance bacteria from 47 strains of isolated E.coli.To analyze correlations between the degree of E.coli aminoglycoside resistance and aac(3) Ⅱgene conserved region,PCR amplified aac(3) Ⅱgene conserved regions and were analyzed by DNA sequencing.The results showed that there were two species of aac(3) Ⅱgene type including 65G and 84T or 65A and 84C in the samples.Strains with high activity of modifying enzyme to gentamicin all were 65G and 84T aac(3) Ⅱgene type.     

Differential and Reciprocal Regulation between Hypoxia-inducible Factor-α Subunits and Their Prolyl Hydroxylases in Pulmonary Arteries of Rat with Hypoxia-induced Hypertension

Hypoxia-inducible factor (HIF)-α subunits (HIF-1α,HIF-2α and HIF-3α),which play a pivotalrole during the development of hypoxia-induced pulmonary hypertension (HPH),are regulated through post-U'anslational hydroxylation by their three prolyl hydroxylase domain-containing proteins (PHD 1,PHD2 and PHD3).PHDs could also be regulated by HIF.But differential and reciprocal regulation between HIF-α and PHDs duringthe development of HPH remains unclear.To investigate this problem,a rat HPH model was established.Meanpulmonary arterial pressure increased significantly after 7 d of hypoxia.Pulmonary artery remodeling indexand right ventricular hypertrophy became evident after 14 d of hypoxia.HIF-1α and HIF-2α mRNA increasedslightly after 7 d of hypoxia,but HIF-3α increased significantly after 3 d of hypoxia.The protein expressionlevels of all three HIF-α were markedly upregulated after exposure to hypoxia.PHD2 mRNA and proteinexpression levels were upregulated after 3 d of hypoxia;PHD 1 protein declined after 14 d of hypoxia withoutsignificant mRNA changes.PHD3 mRNA and protein were markedly upregulated after 3 d of hypoxia,then themRNA remained at a high level,but the protein declined after 14 d of hypoxia.In hypoxic animals,HIF-lotproteins negatively correlated with PHD2 proteins,whereas HIF-2α and HIF-3α proteins showed negativecorrelations with PHD3 and PHD 1 proteins,respectively.All three HIF-α proteins were positively correlatedwith PHD2 and PHD3 mRNA.In the present study,HIF-α subunits and PHDs showed differential andreciprocal regulation,and this might play a key pathogenesis role in hypoxia-induced pulmonary hypertension.     

混合培养真菌和细菌对废水的生物去氮作用

Denitrificationis a biological processin which nitrateand/or nitrite is reduced to gaseous nitrogen,dinitrogen(N2)or nitrous oxide(N2O)while carbon dioxide is thesecond gaseous product of the process.This is one of themain mechanisms of the global nitrogen cycle,and playsanimportant role as the reverse reaction of nitrogen fixa-tion in maintaining global environmental homeostasis[1].Denitrification has beenlongthought to be a unique char-acteristic of prokaryotes[2,3].Anumber of bacteria(such…     

烟草DREBP转录因子结合DRE元件的关键氨基酸

从烟草品种本塞母氏中分离出2条DREB类转录因子基因,分别命名为NbDREB1和 NbDREB2.根据测序结果推导出的氨基酸序列分析显示,NbDREB1和NbDREB2都具有典型的AP2/EREBP转录因子家族EREBP亚族A类特征.酵母单杂交结果显示,它们都不具有激活功能.连接pGADT7反式激活载体形成融合基因表达结果显示,NbDREB1能与DRE顺式作用元件结合,NbDREB2则不能.比较NbDREB1和NbDREB2的AP2区,发现两者的第2和49位氨基酸残基不同.对NbDREB2的第2位氨基酸残基N点突变为Y,NbDREB2也显示出与DRE顺式元件结合的活性,表明烟草DREB转录因子的AP2区第2位氨基酸残基Y是识别及结合DRE顺式作用元件必需的氨基酸残基.     

广藿香八氢番茄红素合成酶PcPSY1基因克隆和序列分析

根据已获得的广藿香转录组数据中的PSY转录本序列,利用Primer 3在线设计基因全长扩增引物,采用RT-PCR方法获得广藿香的八氢番茄红素合成酶(phytoene synthase,PSY)基因。并利用在线分析平台和生物软件对该其因进行生物信息学分析。获得的广藿香PSY1基因长1 550 bp,编码439个氨基酸,命名为PcPSY1,GenBank登录号为KC862310; 预测了PcPSY1 编码蛋白的结构与功能,且基于PSY基因利用NJ法构建了与21个不同物种间的进化树,进化树表明广藿香PcPSY1基因与桂花的PSY序列亲缘关系最近。成功克隆并分析广藿香PcPSY1基因的全长序列,为进一步阐明广藿香萜类代谢途径奠定基础。     

金属螯合亲和层析介质用于六聚组氨酸融合蛋白的纯化研究

以Sepharose CL-6B为载体,环氧氯丙烷为活化剂,羧甲基天冬氨酸(CM-Asp)为螯合配基制备载有Co2 的金属螯合亲和层析介质Co-CM-Asp-Sepharose,并将其用于六聚组氨酸融合蛋白的纯化研究。对纯化200μL细胞裂解液中靶蛋白所需Co-CM-Asp-Sepharose介质用量,Co-CM-Asp-Sepharose与细胞裂解液的孵育时间,介质清洗条件及靶蛋白洗脱时所需咪唑浓度等进行了优化。比较了Co-CM-Asp-Sepharose与Ni-NTA-Agarose(Qiagen公司)两种螯合介质对融合蛋白的纯化效果,开展了从5mL细胞裂解液中放大规模纯化融合蛋白的研究,并通过Bradford法测定了Co-CM-Asp-Sepharose对CD155D1蛋白的纯化量。结果表明:对200μL细胞裂解液纯化体系,Co-CM-Asp-Sepharose(50%悬浮液)的优选体积为60μL,最佳孵育时间为30min,洗脱液最佳咪唑浓度为200mmol/L,纯化得到融合蛋白的量约为200μg。介质用量放大为1.5mL(50%悬浮液)对CD155D1蛋白的纯化量可达4.6mg。与商品化Ni-NTA-Agarose相比,本介质具有选择性好,清洗条件简单,得到的靶蛋白纯度高等优点。     

柑橘大实蝇信息素鉴定及EAG测试

[目的]分析和鉴定柑橘大实蝇Bactocera (Tetradacus) minax (Enderlein)信息素.[方法]采用顶空固相微萃取方法(HS-SPME)收集不同时间段柑橘大实蝇成虫挥发物,进一步结合气相色谱质谱联用技术(GC-MS)鉴定分析和触角电位技术(EAG)进行行为测试.[结果]N-(3-甲基丁基)乙酰胺为柑橘大实蝇成虫合成的一种挥发性化合物,且雌雄成虫都可以释放该化合物.触角电位技术测定不同日龄柑橘大实蝇成虫对1、10、100和1000 μg4种不同剂量N-(3-甲基丁基)乙酰胺的触角电位反应结果表明,柑橘大实蝇成虫对N-(3-甲基丁基)乙酰胺具有显著的触角电位反应,触角电位反应最适含量范围为大于100 μg,剂量以及剂量×性别会导致柑橘大实蝇显著的触角电位反应差异.[结论]N-(3-甲基丁基)乙酰胺可能为柑橘大实蝇信息素,这一结果可为柑橘大实蝇化学通讯研究和生物防治应用提供一定的理论基础.