Location and PCR analysis of catabolic genes in a novel Streptomyces sp. DUT AHX capable of degrading nitrobenzene

A novel strain of Streptomyces sp.DUT_AHX was isolated from sludge contaminated with nitrobenzene and identified on the basis of physiological and biochemical tests and 16S ribosomal DNA (rDNA) sequence analysis.The optimal degradation conditions were as follows:temperature 30℃,pH 7.0-8.0,shaking speed 150-180 r/min,and inocula 10%(V/V).The strain,which possessed a partial reductive pathway with the release of ammonia,was also able to grow on mineral salts basal (MSB) medium plates with 2-aminophenol, ph...     

Isolation of a Pseudomonas Stutzeri strain that degrades 1,2,4-trichlorobenzene and characterization of its degradative plasmid

The genetic information encoding metabolic pathways for xenobiotic compounds in bacteria often resides on catabolic plasmids. The aim of the present work was to know the location of the genes for degrading 1,2,4-trichlorobenzen. In this paper a 1,2,4-trichlorobenzene-degrading strain THSL-1 was isolated from the soil of Tianjin Chemical Plant using 1,2,4-trichlorobenzene as the sole carbon source. The strain was identified as Pseudomonas stutzeri through morphologic survey and 16S rDNA sequence determination. A plasmid was discovered from strain THSL-1 by using the alkali lysis method. When the plasmid was transformed into E. coli. JM109 by the CaCl₂ method, the transformant could grow using 1,2,4-trichlorobenzene as the sole carbon source and had the degradation function of 1,2,4-trichlorobenzene. Therefore, it could be deemed that the plasmid carried the degradative genes of 1,2,4-trichlorobenzene. The average size of the plasmid was finally determined to be 40.2 Kb using selectively three kinds of restricted inscribed enzymes (HindIII, BamHI, and XholI) for single cutting and double cutting the plasmid pTHSL-1, respectively. __________ Translated from China Environmental Science, 2005, 25(4): 385–388 [译自: 中国环境科学]     

Stimulants and donors promote megaplasmid pND6-2 horizontal gene transfer in activated sludge

The activated sludge process is characterized by high microbial density and diversity, both of which facilitate antibiotic resistance gene transfer. Many studies have suggested that antibiotic and non-antibiotic drugs at sub-inhibitory concentrations are major inducers of conjugative gene transfer. The self-transmissible plasmid pND6-2 is one of the endogenous plasmids harbored in Pseudomonas putida ND6, which can trigger the transfer of another co-occurring naphthalene-degrading plasmid pND6-1. Therefore, to illustrate the potential influence of stimulants on conjugative transfer of pND6-2, we evaluated the effects of four antibiotics (ampicillin, gentamycin, kanamycin, and tetracycline) and naphthalene, on the conjugal transfer efficiency of pND6-2 by filter-mating experiment. Our findings demonstrated that all stimulants within an optimal dose promoted conjugative transfer of pND6-2 from Pseudomonas putida GKND6 to P. putida KT2440, with tetracycline being the most effective (100 µg/L and 10 µg/L), as it enhanced pND6-2-mediated intra-genera transfer by approximately one hundred-fold. Subsequently, seven AS reactors were constructed with the addition of donors and different stimulants to further elucidate the conjugative behavior of pND6-2 in natural environment. The stimulants positively affected the conjugal process of pND6-2, while donors reshaped the host abundance in the sludge. This was likely because stimulant addition enhanced the expression levels of conjugation transfer-related genes. Furthermore, Blastocatella and Chitinimonas were identified as the potential receptors of plasmid pND6-2, which was not affected by donor types. These findings demonstrate the positive role of sub-inhibitory stimulant treatment on pND6-2 conjugal transfer and the function of donors in re-shaping the host spectrum of pND6-2.     

质粒pT7降解三氯苯的功能研究

从以1,2,4-三氯苯为唯一碳源的降解菌--施氏假单胞菌(Pseudomonas Stutzeri)T7中提取到一个降解质粒,为研究该质粒的降解功能,将其转化到E.coli JM109中,转化子获得了降解1,2,4-三氯苯的能力.采用色谱/质谱联用(GC/MS)技术检测到转化子E.coli JM109(含质粒pT7)降解1,2,4-三氯苯过程中的部分中间产物分别为2,3,5-三氯-2,4-己二烯二酸、2,5-二氯-2,4-二烯-1,4-内酯-己酸、2,5-二氯-4-氧代-2-烯己二酸;在转化子中分别检测了三氯苯双加氧酶、脱氢酶、邻苯二酚1,2-双加氧酶的酶活性.     

Chemical carcinogenesis — the price for DNA-repair?

This essay examines the possibility of merging the mutation theory of cancer with the hypothesis that cancer is a change in the state of the differentiation of cells. It is suggested that during normal development DNA rearrangements occur, concerning genes which code for differentiation specific cell communication proteins. These proteins are responsible for the proper functioning of growth control in a multicellular organism. DNA-damaging agents — mutagens — induce DNA repair enzymes, some of which may catalyse illegitimate genome rearrangements, thus leading to a change of the balance between growth and differentiation. A cell with a selective advantage may arise and become the origin of a tumor.     

基于玛氏骨条藻(Skeletonema marinoi)转录组的氮代谢途径解析

本研究分别选取了不同生长时期、不同温度培养以及低硅条件培养的玛氏骨条藻（Skeletonema marinoi）,以Illumina Hiseq 2000平台进行转录组高通量测序分析,共获得39,098个转录本。通过NR注释、GO注释和KEGG通路注释等一系列生物信息学手段对该转录组进行了基因注释和代谢途径分析。在此基础上重点分析了玛氏骨条藻的氮代谢途径,发现共存在20种酶和蛋白及6条相关代谢途径。对38个编码这些酶的基因序列比对结果表明,其与假微型海链藻同源基因具有较高的一致性。同时对这些样品进行数字基因表达谱分析,获得不同生长时期氮代谢途径中酶和蛋白的编码基因的差异表达情况:与对数期相比,在稳定期参与硝酸盐同化过程的两种酶（硝酸盐还原酶、亚硝酸还原酶）的基因和参与氨氮代谢的两种酶（氨甲酰磷酸合成酶和氨基甲酸激酶）的基因均有明显的上调表达,这是玛氏骨条藻对所处的不同生长状态下氮元素存在形态的变化所做出的分子响应。由转录组测序构建的代谢途径及相关基因的差异表达信息有助于对玛氏骨条藻氮代谢关键酶基因调控过程的解析,为进一步研究赤潮藻氮素营养限制过程中相关基因的表达调控模式奠定了基础,可为深入了解赤潮藻对营养元素的利用提供依据。     

蚯蚓金属硫蛋白定量PCR检测方法及其分子诊断

根据GenBank中蚯蚓(Eisenia fetida)金属硫蛋白(MT)基因序列设计合成引物,采用RT-PCR扩增出目的基因片段,将目的基因片段插入pEASY-Blunt中制备质粒标准品.通过不同梯度稀释倍数的质粒标准品,建立了目的基因MT与内参基因β-actin的SYBR Green I荧光定量RT-PCR检测方法.通过土壤染毒培养实验,将蚯蚓分别暴露于50,500,1000mg/kg 重金属Cd染毒的自然土壤中培养14,28d,研究蚯蚓体内MT mRNA转录水平的表达变化.结果表明,重组质粒测序后显示目的片段序列同源性好,证明所设计的MT、β-actin引物能成功用于蚯蚓MT mRNA的检测.两基因构建的荧光定量标准曲线线性关系分别为0.994,0.999,且PCR扩增效率也皆接近于100%.染毒实验发现,Cd可诱导蚯蚓体内MT mRNA的表达,其表达水平与Cd污染暴露呈现出剂量-效应和时间-效应关系,表明蚯蚓MT基因可作为潜在分子标志物,诊断环境中Cd污染及其暴露水平.     

Biosensors: new approaches in drug discovery

The development of biosensors for analytical purposes has attracted a great deal of attention in recent years. A biosensor is defined as an analytical device consisting of a biological component (e.g., enzyme, antibody, entire cell, DNA) and a physical transducer (e.g., electrode, optical device). Biosensors are mostly designed for routine analysis, such as clinical diagnosis, quality control of food, in-process control of fermentations, and in environmental analysis. Many of these sensors are also suitable for screening purposes in order to find new drugs. Such systems should yield information either about compounds with known bioactivity or about the bioactivity of samples with known or unknown chemical composition. Biosensors intended for the latter purpose are essentially based on whole cells carrying receptors and ion channels at their surfaces. Miniaturization of structures, primarily based on silicon, allows integration of many sensors into arrays, which may be suitable for the screening of natural and chemical products as well as combinatorial libraries. Until now, no commercially available sensors of this kind exist but they are expected in the near future. Different biosensors, based on enzymes, antibodies, cells, artificial membranes and entire animal tissues, which can be used in drug discovery and may lead to efficient screening systems in the future, are described in this review.     

零价铁对污泥厌氧消化过程中四环素抗性基因水平转移的作用影响

细菌抗药性问题已成为全球生态环境安全和人体健康领域的一个关注焦点.污水处理剩余污泥是抗生素抗性基因（ARGs）的一个重要储存库和排放源,采用荧光定量PCR方法检测分析了四环素抗性基因（TC-ARGs,包括tetA、tetC、tetG、tetM、tetO、tetW和tetX）及第一类整合子 intI1基因丰度在污泥厌氧消化过程中的变化特征,考察了不同投加量零价铁（Fe⁰）对目标基因的消长影响,初步分析了质粒DNA接合作用对TC-ARGs水平转移的作用影响,并探讨了TC-ARGs与 intI1基因之间的相关关系.结果表明,污泥厌氧消化过程对TC-ARGs和intI1基因丰度具有不同程度的削减作用,其中tetX基因丰度降幅可达2.4个数量级.Fe⁰的加入对TC-ARGs和intI1基因丰度削减不显著,并且随着Fe⁰投加量的增加,TC-ARGs和intI1基因丰度相比对照组出现逐渐升高的趋势.通过对质粒组DNA所携带的TC-ARGs基因丰度进行检测分析发现,质粒DNA接合作用对TC-ARGs的水平转移具有促进作用.相关性分析结果显示,tetG与intI1基因之间具有显著正相关关系,表明intI1基因在污泥厌氧消化过程中可能对tetG基因的演变过程具有重要影响.     

Occurrence,functions and biosynthesis of polyamides in microorganisms and biotechnological production

Microorganisms are able to synthesize three different polyamides by enzymatic processes independently from ribosomal protein biosynthesis: poly(gamma-D-glutamic acid), poly(epsilon-L-lysine) and multi-L-arginyl-poly(L-aspartic acid) which is also referred to as cyanophycin. These polyamides, which occur mainly in Bacillus spp. (and only a few other eubacteria and the nematocysts of Cnidaria, in Streptomyces albulus or in cyanobacteria, respectively), have recently attracted considerable interest of the chemical industry and may be suitable for various applications. This review summarizes our current knowledge on the occurrence, biosynthesis, physiological functions, and biodegradation as well as on the properties and putative applications of these polyamides. Emphasis is placed on the enzymology of the polymerization and on the genes encoding the polymerizing enzymes, which have only recently become available for cyanophycin synthetases. Prospects for novel production processes. in particular for cyanophycin, are also presented.     

Zentralnervöse Sphingolipid-Speicherkrankheiten

Lysosomal storage disorders are progredient and often fatal diseases most of which result from a pronounced enzyme deficiency. In the case of sphingolipidoses, usually enzymes of sphingolipid catabolism are missing, or only a few percent of normal activity are detectable. For many sphingolipidoses, damage of the central nervous system is characteristic, but neurological and other symptoms can vary greatly, especially in adult variants. This variability is mainly caused by different allelic mutations of the structural genes, resulting in different levels of residual enzyme activity.     

城市污水中病原性大肠埃希氏菌毒力基因eaeA和rfbE的实时荧光定量PCR检测

针对病原性大肠埃希氏菌EHEC和EPEC的毒力基因eaeA和rfbE,选用特异性引物,建立了实时荧光定量PCR检测方法. 利用从污水中分离出的目的核酸片段构建重组质粒,通过测序和BLAST比对分析,确定了PCR扩增的特异性. 将重组质粒作为模板,分别测定标准曲线,在eaeA基因模板量8.77～8.77×105 copy,rfbE基因模板量4.98～4.98×105copy的范围内,Ct(循环阈值)与模板量的对数值具有良好的线性关系〔R2为0.997(eaeA)和1.000(rfbE)〕. 结合膜吸附洗脱浓缩方法,对于实际水样,eaeA和rfbE基因的检测限分别为1.75×102和9.96×101copy/100 mL. 利用该方法对城市污水处理厂进、出水进行检测的结果显示,污水二级处理工艺对这2种毒力基因的去除效果明显.      

Transportkatalyse in Biomembranen

A basic issue of biomembranes is their ability to facilitate specific transport of selected molecules. This transport is catalyzed by carriers which are membrane proteins and form, analogous to enzymes, carrier-substrate complexes. The ADP,ATP carrier of mitochondria is highly suitable for elucidating the mechanism of this catalysis due to its unique qualities such as great abundance in higher cells, easy isolation in native state by detergents, existence of inhibitors specific for either the in- or outward looking binding site and direct observation of a carrier-substrate complex. As central catalytic steps, the reorientation of the substrate-binding site at the carrier during translocation across the membrane could be demonstrated at the intact membrane and the isolated protein. The results are interpreted by the gated-pore mechanism where two subunits form a gate with a central binding site which radically change conformation and specificity on transition from one to the other side of the membrane.     

诺卡氏菌株C-14-1中腈水解酶基因的鉴定、测序及分析

从腈纶废水中分离到高效降解多种污染物的诺卡氏菌株C-14—1,并对该菌中腈水解酶的基因进行鉴定和测序．利用红球菌中腈水解酶氨基酸保守区设计核苷酸引物,以菌株C-14—1的总DNA为模板,PCR扩增发现一条预期大小的DNA带．Southern杂交显示基因组中存在一个腈水解酶基因．进一步构建基因文库和菌落原位杂交,克隆到一个约4．5kb的DNA片段．DNA序列测定和分析表明,该DNA片段携带长度为1143bp的腈水解酶基因．比对分析表明,该基因与国际上发表的红球菌和诺卡氏菌中的腈水解酶基因高度相似．     

工业循环冷却水系统阻垢剂研究进展

本文简介了循环冷却水系统的应用现状和面临的问题．同时论述了冷却水系统结垢的成因和阻垢的机理,并介绍了阻垢剂的发展历程、研究进展及其特点。最后总结概括了未来阻垢剂的发展趋势。     

H2S信号在拟南芥响应SO2胁迫过程中的作用

以拟南芥为实验材料,研究植物对H₂S和SO₂处理的转录响应及其关系,并利用外源喷施H₂S及其清除剂的方法,检测SO₂熏气后植株体内H₂S的产生及其生理效应,探讨气体信号分子H₂S在植物响应SO₂胁迫过程中的作用.高通量测序结果发现,H₂S和SO₂处理诱导拟南芥植株多个基因转录水平改变,并有1220个基因在两种处理条件下均差异表达,其中包括多个硫代谢和谷胱甘肽代谢相关基因,表明H₂S和SO₂在调控硫代谢途径中具有交互作用.SO₂熏气诱导拟南芥植株体内H₂S合成酶基因LCD和DES1转录水平提高,胞内H₂S含量增多.同时,超氧化物歧化酶(SOD)和过氧化氢酶(CAT)活性升高,含硫抗氧化物谷胱甘肽(GSH)含量及其相关酶谷胱甘肽硫转移酶(GST)和谷胱甘肽过氧化物酶(GPX)活性提高,活性氧H₂O₂含量增加,膜脂过氧化产物丙二醛(MDA)含量与对照相比无显著差异.外源喷施H₂S可进一步提高SO₂熏气下拟南芥植株GSH含量及其相关防御酶的活性,H₂O₂含量降低.但喷施亚牛磺酸(HT)清除H₂S后,SO₂熏气拟南芥植株GSH含量、抗氧化酶SOD、CAT和GST活性降低,MDA含量大幅增加.结果表明,SO₂熏气诱导产生的H₂S可作为信号分子,提高机体抗氧化防御能力,增强植物对SO₂胁迫的抗性.     

假单胞菌的氯儿茶酚1,2-双加氧酶基因的克隆和表达

 从土壤中分离得到1株降解2,4-二氯酚(2,4-DCP)能力较强的细菌菌株GT241-1,克隆了该菌株的3,5-二氯儿茶酚1,2-双加氧酶基因(dcpB),采用的克隆策略为:用Southern杂交对dcpB进行定位后,构建重组质粒,再用斑点杂交从重组质粒中筛选目的转化子.经序列测定得知dcpB亚克隆片段全长4303bp,其中dcpB基因编码区765bp.核苷酸和推测的氨基酸序列分析表明,dcpB与已在GenBank登记的相关基因有一定的差异.dcpB基因能够在大肠杆菌转化子中成功地表达有生物活性的酶.     

水产致病菌耐药基因的研究

采用纸片扩散法(Kirby-Bauer法)研究不同来源的54株水产细菌对10种抗生素的敏感性,筛选获得耐药菌株并研究其耐药基因。根据GenBank中注册的耐药基因序列,针对不同抗生素的耐药基因设计相应的引物,对耐药基因进行扩增,检测耐药菌株中耐药基因的分布。结果由54个受试菌株中检测出42个耐药株,耐药率为77.8%。其中37株对3种以上抗生素产生耐药,多重耐药率为68.5%。以耐药菌株质粒为模板,扩增磺胺类耐药基因sul2、氯霉素耐药基因cat1、cat2、cat3、cat4、卡那霉素耐药基因aadB、喹诺酮耐药基因gyrA,结果显示:sul2基因阳性9株,3株氯霉素cat2阳性,4株氯霉素cat3阳性,1株氯霉素cat4阳性,3株aadB阳性。质粒上耐药基因的检出率分别为50%、27%、36%、9%、60%。     

“大数据”预测分析与三丁基锡暴露有关疾病

通过毒理学“大数据”,挖掘三丁基锡（TBT）潜在的生物影响信息,预测分析TBT暴露与人类相关疾病的关系.通过比较毒理组学数据库（CTD）搜集到488个和TBT具有相互作用的基因.基因相互作用网络图显示TP53拥有相关联基因最多,处于中心地位;其次是ESR1和FN1.CTD分析与TBT相关的疾病,发现前10类疾病分别是癌症、神经系统疾病、心血管疾病、泌尿生殖系统疾病（女）、消化系统疾病、代谢疾病、泌尿生殖系统疾病（男）、内分泌系统疾病、免疫系统疾病、呼吸道疾病.KEGG通路和DAVID基因功能注释分析,发现TBT不仅和代谢性疾病有较高的关联性,其相互作用基因在糖代谢途径上亦有较密集的基因功能注释.PASS的活性预测也发现TBT可能影响多种与糖代谢相关的酶生物活性.这提示TBT对糖代谢相关疾病的影响应引起人们的注意.