非常规酵母的分子遗传学及合成生物学研究进展

先进的合成生物学技术与传统的分子遗传学技术的结合更有助于实现酵母底盘细胞的快速改造和优化。酵母合成生物学研究最早开始于常规酵母——酿酒酵母(Saccharomyces cerevisiae),近些年来又迅速扩展至一些非常规酵母,包括巴斯德毕赤酵母(Pichiapastoris)、解脂耶氏酵母(Yarrowialipolytica)、乳酸克鲁维酵母(Kluyveromyces lactis)和多形汉逊酵母(Hansenula polymorpha)等。借助合成生物学技术与工具,目前科学家们已经成功开发出了能够高效生产生物材料、生物燃料、生物基化学品、蛋白质制剂、食品添加剂和药物等工业产品的重组非常规酵母工程菌株。本文系统总结了合成生物学工具(主要是基因组编辑工具)、合成生物学组件(主要是启动子和终止子)和相关分子遗传学方法在上述非常规酵母系统(底盘细胞)中的最新研究进展和应用情况,并讨论了其他合成生物学技术在这些非常规酵母表达系统中的潜在适用性和应用前景。这为研究人员利用合成生物学方法在这一新型非模式微生物底盘细胞中设计和构建各种高附加值工业产品的异源合成模块并最终实现目标化合物的高效生物合成提供了科学的理论指导。     

《实验生物学与医学》：科学家研发出胞内即时影像检测共振传感器

来自EurekAlert的消息。伊利诺大学基因组生物学研究所的研究人员突破了FRET技术的局限性，开发了一个共振传感器。来检测细胞内的氧化还原动力学的即时影像，这一研究成果公布在《实验生物学与医学》（Experimental Biologyand Medicine）杂志上。     

合成生物学中的正交遗传系统

并行、独立的正交系统是合成生物学的重要研究基础之一,这个系统与自然界的生物系统及其组成交叉很少或没有交叉。它的组成包括非天然碱基对、移位密码子、非天然氨基酸、正交的氨酰tRNA合成酶、RNA聚合酶和启动子、正交核糖体等。这些正交系统的组成部分可以一起组成系统发挥作用,也可以各自单独在生物体系中应用,它们给生物带来新的特性,也为研究人员提供了新的生物学研究方法。     

儿茶素EGCG与其空间异构体GCG的抗氧化活性和反应活性的密度泛函理论研究

为了分析儿茶素EGCG及其空间异构体GCG自由基清除活性的差异,在6-311++G(d,p)水平下采用密度泛函理论中的B3LYP方法计算EGCG和GCG的酚羟基键解离能(BDE)、电离能(IP)、电子亲和势(EA)、EHOMO和ELUMO能级及其能级差Egap等性质。结果表明:EGCG和GCG的自由基清除反应最有可能发生在B环,这主要是由于B环上的相邻酚羟基之间的弱氢键作用所产生的共轭稳定性;此外研究还发现,GCG比EGCG稳定,但两者具有相同的基于氢原子转移机制(HAT)和电子转移随后发生质子迁移机制(SPLET)的自由基清除活性。     

《实验生物学与医学》：科学家研发出胞内即时影像检测共振传感器

来自EurekAlert的消息，伊利诺大学基因组生物学研究所的研究人员突破了FRET技术的局限性，开发了一个共振传感器，来检测细胞内的氧化还原动力学的即时影像，这一研究成果公布在《实验生物学与医学》（Experimental Biology and Medicine）杂志上。     

合成生物学助力应急疫苗研制

合成生物学具有系统性思维和工程学理念的特点,致力于创造新的生命或新的系统。合成生物学为疫苗研发人员提供了新的思路,不断开发出新疫苗研制的技术平台。通过抗原重构技术,合成的强抗原能够很好地激活VRC0-1种系的B细胞;利用合成生物学合成自组装的纳米颗粒疫苗,能够有效地暴露保守抗原;通过合成生物学完成流感疫苗基因组的快速组装,大大缩短了疫苗的研制周期;通过系统性地改造,重组沙门菌再一次成为疫苗开发的重要工具。合成生物学在应急疫苗研发过程中疫苗研制平台搭建及快速合成新发传染病抗原方面发挥着突出作用。     

合成生物学在基础生命科学研究中的应用

合成生物学作为一门新兴的交叉学科,吸引了来自生物学、数理科学和工程学等不同学科的研究人员以及产业界的广泛关注和参与。它旨在通过从头创造全新的或改造已有的生物系统,实现天然生物系统不具备的功能与特性。合成生物学研究不仅具有广阔的生物产业应用前景,更为基础科研提供了全新的手段和思路。本文着眼于合成生物学―建物致知‖的理念,跟踪合成生物学研究在回答生命科学基础问题方面取得的相关成果,简述了其在细胞内分子调控网络、细胞生理学、多细胞群体形态与行为以及多物种微生态学等研究中的应用。     

合成生物学发展现状与前景

合成生物学是综合了科学与工程的一个崭新的生物学研究领域,为生命现象及其运动规律的解析提供了一种采用“白下而上”合成策略的正向工程学的研究思路和方法手段,在经济和社会发展中具有巨大的应用开发潜力。近年来,DNA合成与系统生物学技术的发展使生命系统复杂基因回路的设计、合成与组装逐步成为可能,并应用于生物基化学品、生物燃料、医药中间体、保健产品的生产和环境保护等领域。但是,合成生物学的研究仍然面临科学、技术和伦理的挑战,只有积极地应对这些问题,在加大研究开发支持力度的同时,做好必要的风险监管,才能真正把握合成生物学发展带来的历史机遇。     

毛丝鼠在医学研究中的应用

毛丝鼠已成功用于听觉系统、微生物及寄生虫感染动物模型研究,由于其独特的生物学特点,可进一步开发以用于老年性疾病、代谢性疾病等方面研究。本文将介绍毛丝鼠的相关生物学特性,并就其在医学研究中的应用进展作简要综述。     

合成生物学指导下芽孢杆菌合成环脂肽的研究进展*

近年来,合成生物学借助工程化在人工生命系统的设计与构建方面取得了长足进展,特别是“细胞工厂”的开发和应用为天然产物的合成带来了深刻变革。环脂肽是一类新型的天然表面活性剂,因其特殊的结构和功能亦可作为抗生素使用。目前,合成环脂肽最理想的微生物底盘是芽孢杆菌。因此,许多研究者致力于通过合成生物学技术来提升芽孢杆菌作为环脂肽细胞工厂的性能。首先,对芽孢杆菌中环脂肽的非核糖体肽合成途径进行概述;其次,重点介绍与环脂肽合成相关的调控因子;再次,从底盘细胞的选择、基因编辑工具的开发、合成路径的优化及发酵过程的优化等四个方面对合成生物学指导下环脂肽的相关研究进展进行总结;最后,讨论环脂肽合成中可能存在的挑战,并就未来研究趋势进行展望,以期为高效环脂肽细胞工厂的开发提供参考。     

Localization of Escherichia coli RNA polymerase binding sites on bacteriophage S13 and 0X174 DNAs by electron microscopy.

Complexes between Escherichia coli RNA polymerase and bacteriophage S13 and phage phiX174 replicative form III DNAs have been shown to form at specific locations on the phage genomes. The major locations on S13 have been mapped at 8 to 10 and 92 to 96% of the genome length, starting from the unique Pst I cleavage site. The locations correspond to the beginnings of genes D and B, respectively. Four minor locations map at 18 to 22, 28 to 32, 50 to 56, and 70 to 74% of the genome. The 70 to 74% site corresponds to the beginning of the A gene. The major locations on phiX174 are at 8 to 10, 50 to 54, and 92 to 94% of the genome. The 50 to 54% site is at the start of the H gene and has an equivalent minor site on S13, but it is not a promoter site. Three minor sites on phiX174, at 20 to 24, 26 to 32, and 68 to 74% of the genome, correspond to sites on S13. The data confirm the locations of sites identified by restriction fragment binding experiments (E. Rassart and J. H. Spencer, J. Virol. 27:677--687, 1978) and the assignment of putative promoters at the start of genes A, B and D.     

西双版纳热带雨林蚁科昆虫区系分析

在西双版纳热带雨林已鉴定蚊科昆虫９亚科７６属２６７种。西双版纳地区的蚂蚁区系以热带至亚热带分布的东洋界成分最为丰富。在属级水平上,与马来西亚界关系最为密切。与澳洲界关系较密切;与非洲界和马拉加西界的关系知中。与新北界,新热带界和古北界的关系最为疏远。可见西双版纳的蚂蚁区系具有典型的热带亚洲起源特征,同时与澳洲和非洲的热带区系有一定的渊源关系。     

Specificity of the binding of trifluoperazine to the calcium-dependent activator of phosphodiesterase and to a series of other calcium-binding proteins

Trifluoperazine inhibits the activation of phosphodiesterase by binding to the calcium-dependent activator. To determine further the specificity by which trifluoperazine binds to activator, we compared the binding of trifluoperazine to activator prepared from several species and tissues and to a number of other calcium-binding proteins devoid of activator activity.Trifluoperazine binds to activator prepared from human, bovine, rat and rabbit brain and from chick embryo fibroblasts. In each case, the binding of trifluoperazine to activator was qualitatively similar and related quantitatively to the ability of the preparation to activate phosphodiesterase.Of the other calcium-binding proteins examined, namely, troponin-C, S-100 protein, phospholipase A, phospholipase B and myosin light chain, only troponin-C displayed any significant calcium-specific binding of trifluoperazine. The binding to troponin-C, however, appeared to be different from the binding to activator; whereas the binding of trifluoperazine to actovator showed no cooperativity, the binding to troponin-C showed positive cooperatively.These results and earlier data showing that trifluoperazine fails to bind to a variety of other proteins, indicate that the binding of trifluoperazine to the calcium-dependent activator of phosphodiesterase is selective and suggest that this binding may explain some of the biochemical and pharmacological actions of this antipsychotic agent.     

Role of the adaptor protein CIKS in the activation of the IKK complex

Nuclear factor kappaB (NF-kappaB) plays a pivotal role in numerous cellular processes, including stress response, inflammation, and protection from apoptosis. Therefore, the activity of NF-kappaB needs to be tightly regulated. We have previously identified a novel gene, named CIKS (connection to IkappaB-kinase and SAPK), able to bind the regulatory sub-unit NEMO/IKKgamma and to activate NF-kappaB. Here, we demonstrate that CIKS forms homo-oligomers, interacts with NEMO/IKKgamma, and is recruited to the IKK-complex upon cell stimulation. In addition, we identified the regions of CIKS responsible for these functions. We found that the ability of CIKS to oligomerize, and to be recruited to the IKK-complex is not sufficient to activate the NF-kappaB. In fact, a deletion mutant of CIKS able to oligomerize, to interact with NEMO/IKKgamma, and to be recruited to the IKK-complex does not activate NF-kappaB, suggesting that CIKS needs a second level of regulation to efficiently activate NF-kappaB.     

Surface topology of the Escherichia coli K-12 ferric enterobactin receptor.

Monoclonal antibodies (MAb) were raised to the Escherichia coli K-12 ferric enterobactin receptor, FepA, and used to identify regions of the polypeptide that are involved in interaction with its ligands ferric enterobactin and colicins B and D. A total of 11 distinct FepA epitopes were identified. The locations of these epitopes within the primary sequence of FepA were mapped by screening MAb against a library of FepA::PhoA fusion proteins, a FepA deletion mutant, and proteolytically modified FepA. These experiments localized the 11 epitopes to seven different regions within the FepA polypeptide, including residues 2 to 24, 27 to 37, 100 to 178, 204 to 227, 258 to 290, 290 to 339, and 382 to 400 of the mature protein. Cell surface-exposed epitopes of FepA were identified and discriminated by cytofluorimetry and by the ability of MAb that recognize them to block the interaction of FepA with its ligands. Seven surface epitopes were defined, including one each in regions 27 to 37, 204 to 227, and 258 to 290 and two each in regions 290 to 339 and 382 to 400. One of these, within region 290 to 339, was recognized by MAb in bacteria containing intact (rfa+) lipopolysaccharide (LPS); all other surface epitopes were susceptible to MAb binding only in a strain containing a truncated (rfaD) LPS core, suggesting that they are physically shielded by E. coli K-12 LPS core sugars. Antibody binding to FepA surface epitopes within region 290 to 339 or 382 to 400 inhibited killing by colicin B or D and the uptake of ferric enterobactin. In addition to the FepA-specific MAb, antibodies that recognized other outer membrane components, including Cir, OmpA, TonA, and LPS, were identified. Immunochemical and biochemical characterization of the surface structures of FepA and analysis of its hydrophobicity and amphilicity were used to generate a model of the ferric enterobactin receptor's transmembrane strands, surface peptides, and ligand-binding domains.     

Specificity of the binding of trifluoperazine to the calcium-dependent activator of phosphodiesterase and to a series of other calcium-binding proteins.

Trifluoperazine inhibits the activation of phosphodiesterase by binding to the calcium-dependent activator. To determine further the specificity by which trifluoperazine binds to activator, we compared the binding of trifluoperazine to activator prepared from several species and tissues and to a number of other calcium-binding proteins devoid of activator activity. Trifluoperazine binds to activator prepared from human, bovine, rat and rabbit brain and from chick embryo fibroblasts. In each case, the binding of trifluoperazine to activator was qualitatively similar and related quantitatively to the ability of the preparation to activate phosphodiesterase. Of the other calcium-binding proteins examined, namely, troponin-C, S-100 protein, phospholipase A, phospholipase B and myosin light chain, only troponin-C displayed any significant calcium-specific binding of trifluoperazine. The binding to troponin-C, however, appeared to be different from the binding to activator; whereas the binding of trifluoperazine to actovator showed no cooperativity, the binding to troponin-C showed positive cooperatively. These results and earlier data showing that trifluoperazine fails to bind to a variety of other proteins, indicate that the binding of trifluoperazine to the calcium-dependent activator of phosphodiesterase is selective and suggest that this binding may explain some of the biochemical and pharmacological actions of this antipsychotic agent.     

细菌抵抗消毒剂及其对抗生素共耐药

消毒剂是一种可杀灭物体表面、器材设备、皮肤、空气和水源等传播媒介上携带的病原微生物的有机分子。它在体外能杀灭病原微生物,切断其传播途径,进而达到控制污染的目的,在生命安全防控中起着重要的作用。但是不合理地使用消毒剂导致细菌对消毒剂产生耐药。消毒剂耐药基因在不同种属间的水平转移加剧其传播风险,使消毒剂耐药情况进一步恶化。更令人担忧的是,细菌对消毒剂的耐药可能会导致对抗生素产生共耐药,给公共安全带来巨大的威胁。但目前为止,对消毒剂耐药以及共耐药的认识还不够全面。本文总结了关于细菌对消毒剂耐药的研究报道,对消毒剂的作用机制、细菌对消毒剂的耐药机制进行了论述,另外针对消毒剂耐药基因的传播以及细菌对消毒剂和抗生素的共耐药进行了综述,为减少消毒剂耐药性的产生和制定合理的消毒剂使用规范奠定基础。     

Quantitative data on training new world primates to urinate

This study assessed the effectiveness of operant conditioning in training three species of captive callitrichid primates (Leontopithecus rosalia, Callithrix geoffroyi, and Saguinus imperator) to urinate on demand. There were three goals to the study: 1) to develop a system for quantitatively assessing positive reinforcement training; 2) to ascertain whether or not positive reinforcement techniques can be used to train callitrichid monkeys to urinate on demand, and if so, how many training sessions are required; and 3) to determine the effect on urination behavior of the trainer entering the cage to collect a urine sample. Positive reinforcement with a continuous reinforcement schedule was used to capture a natural behavior: urination. Training sessions (30 min each) were conducted at dawn thrice weekly during five consecutive phases: habituation, control, training (animals were rewarded for urinating), maintenance (animals had reached a defined training criteria and continued to be rewarded for urinating), and collection (animals were rewarded for urinating, and the trainer entered the cage to collect the sample). The numbers of 30-min training sessions required to train the three monkey species (L. rosalia, C. geoffroyi, and S. imperator) were five, six, and eight, respectively. For the three species, the mean number of urinations per animal was significantly greater during the training, maintenance, and collection phases compared to the control phase. However, the three species differed significantly in the manner in which the rates of urination changed across the five phases. A higher proportion of subjects urinated during the training, maintenance, and collection phases compared to the control phase. Latency to first urination varied significantly across the five phases, with significantly reduced latencies to urinate during the training, maintenance, and collection phases compared to the control phase. The entry of the trainer into the cage to collect the urine sample did not appear to alter urination behavior. We demonstrate that operant conditioning techniques, which typically incur minimal cost, time investment, and disturbance, can be used to increase the quantity of urine samples collected for physiological analysis, the proportion of animals that urinate, and the speed of sample collection.     

宁波沿海平原Z02孔第四纪地层及古环境演变

宁波沿海平原第四纪沉积物属于海陆交互沉积,记录了良好的古气候变化信息,是研究古气候冷暖变化、沉积物沉积特征的良好载体.对位于宁波沿海平原东南部的Z02孔进行14C和古地磁测年,根据钻孔岩性、孢粉组合、有孔类和介形类组合、粒度等环境替代指标特征,确定了 Z02孔第四纪地层划分,揭示了研究区第四纪以来的古环境演变信息.结果...     

Fluorescence location of RVLM kainate microinjections that alter the control of breathing

Kainic acid (4.7 mM) applied to the rostral ventrolateral medulla (RVLM) surface decreases phrenic output, CO2 sensitivity, and blood pressure in chloralose-urethan-anesthetized, vagotomized, paralyzed, glomectomized, servoventilated cats. In this study using the same preparation, bilateral 50- to 100-nl kainate injections just below the RVLM surface better localized these responses topographically. The physiological responses to unilateral 10-nl kainate injections were then correlated with anatomic location determined by fluorescent microbeads (0.5 micron diam). Many sites were associated with no effect, a few rostral and caudal sites with increased phrenic activity, and cluster of sites with decreased phrenic activity often to apnea, decreased CO2 sensitivity, and decreased responses to carotid sinus nerve stimulation. Blood pressure was unaffected. These sites, within 400 microns of the surface, were ventral to the facial nucleus, ventrolateral to the nucleus paragigantocellularis lateralis, caudal to the superior olive, and rostral to the retrofacial nucleus. They appeared to be within the recently described retrotrapezoid nucleus, which contains cells with respiratory-related activity and projections to the dorsal and ventral respiratory groups. Cells within this site appear able to provide tonic input to respiration and to affect peripheral and central chemoreception.