溶葡球菌酶高效表达与应用

溶葡球菌酶(lysostaphin,Lys)是采用基因克隆技术使溶葡球菌酶基因实现外源表达所产生的蛋白质。它是Zn2+依赖的金属蛋白酶,具有肽链内切酶活性,能专一性地水解葡萄球菌细胞壁Gly五肽桥联,使金黄色葡萄球菌(特别是MRSA)细胞壁破裂,达到溶菌杀菌作用,而不产生耐药性。作为一种抗菌剂,在兽药与临床等领域具有巨大的应用潜力。综述对溶葡球菌酶的来源、作用机制、不同表达系统及前景与展望进行综述。     

An efficient method for stochastic simulation of biological populations in continuous time

We present an efficient algorithm for individual-based, stochastic simulation of biological populations in continuous time. A simple method for its implementation is given and it is compared to Gillespie's commonly used Direct Method. These two methods are proven to be exactly equivalent and, using a basic evolutionary model, it is demonstrated that the new algorithm can run thousands of times faster. Furthermore, while computational cost per event increases linearly with population size under the Direct Method, this cost is independent of population size under the new algorithm. We argue that this gain in efficiency opens up the possibility to explore a new class of models in population biology.     

Efficient shRNA delivery into B and T lymphoma cells using lentiviral vector-mediated transfer

RNA interference is a powerful tool for the functional analysis of proteins by specific gene knockdown. In this study, we devised a rapid and efficient way to screen suitable siRNA sequences and subsequently employ them for specific gene knockdown in usually hard-to-transfect lymphoid cell lines, using a self-inactivating lentiviral vector. Two proteins with different half-lives were chosen, cyclin D1 and STAT3. A specific lacZ reporter fusion assay was used to identify highly effective siRNA sequences. Only siRNA molecules with more than 85% of knockdown efficiency were selected for the generation of lentiviral transfer vectors. Transduction rates of 75–99% were achieved in the lymphoma cell lines Granta 519 (mantle cell lymphoma), Karpas 299, and SUDHL-1 (anaplastic large T cell lymphoma), as demonstrated by green fluorescent protein expression in fluorescence-activated cell sorting analysis. The high level of transduction efficiency allows RNA interference studies to be performed on transduced cells without further manipulation, such as cell sorting or cloning. The LacZ reporter system together with the lentivirus technology is a very important tool in the hematology field, which enables experiments in lymphoid cells that were not possible before.

Electronic supplementary material

The online version of this article (doi:10.1007/s12308-008-0020-x) contains supplementary material, which is available to authorized users.     

A new semiparametric estimation method for accelerated hazard model

The accelerated hazard model has been proposed for more than a decade. However, its application is still very limited, partly due to the complexity of the existing semiparametric estimation method. We propose a new semiparametric estimation method based on a kernel-smoothed approximation to the limit of a profile likelihood function of the model. The method leads to smooth estimating equations and is easy to use. The estimates from the method are proved to be consistent and asymptotically normal. Our numerical study shows that the new method is more efficient than the existing method. The proposed method is employed to reanalyze the data from a brain tumor treatment study.     

Landscape structure versus the effectiveness of nature conservation: Roztocze region case study (Poland)

The objective of the study was to examine the extent to which landscape metrics could be used as an indicator of efficient management of protection areas. The selected sampling areas were natural–landscape units distinguished within the Roztocze Region located in the central-east Poland. Among 446 units, those representing three typological groups determined by three factors were selected. The factors included (1) the area of the unit; (2) the dominant or characteristic type of ecosystem; and (3) the form of protection. Subsequently, thirteen landscape metrics were calculated with the application of the Fragstats software. The analysis revealed high correlation between the form of nature protection and the majority of the calculated indexes. National park units show the highest landscape diversity and stability of various types of ecosystems. This suggests high conservation efficiency. Landscape park units are distinguished by fragmented patchy composition, and spatial structure even less stable than that of the non-protected areas. The study results show that landscape metrics could be used as an indicator of efficient management of different forms of nature protection. They provide an insight into the structure and functioning of the environment at various levels of its organisation. We particularly found diversity metrics to be useful for indicating whether nature conservation goals are archived, and the size and density metric for measuring human interference in the landscape. The selection and interpretation of indexes should be determined by the specific character of a given area.     

Caulerpenyne from Caulerpa taxifolia: A comparative study between CPC and classical chromatographic techniques

Caulerpenyne (Cyn) is a cytotoxic compound firstly isolated in 1978 from Caulerpa prolifera. This metabolite, constituted by a highly reactive diacetoxybutadiene moiety, exhibited a wide range of biological properties with mainly antibacterial properties and antitumoral activities. Few structure–activity relationships (SAR) are available to design more potent bioactive derivatives by pharmacomodulation. Cyn can be produced by total synthesis or extracted from natural sources in particular the green alga Caulerpa taxifolia. Since conventional chromatographic procedures to isolate Cyn from C. taxifolia are time- and solvent-consuming, it was crucial to find a more efficient process to obtain pure Cyn. In our study, Cyn has been purified from C. taxifolia with two different techniques: Centrifugal partition chromatography (CPC) and a classical chromatographic process. The comparative study showed that CPC constitutes a very simple and efficient process to access Cyn.     

HomBlocks: A multiple-alignment construction pipeline for organelle phylogenomics based on locally collinear block searching

Organelle phylogenomic analysis requires precisely constructed multi-gene alignment matrices concatenated by pre-aligned single gene datasets. For non-bioinformaticians, it can take days to weeks to manually create high-quality multi-gene alignments comprising tens or hundreds of homologous genes. Here, we describe a new and highly efficient pipeline, HomBlocks, which uses a homologous block searching method to construct multiple sequence alignment. This approach can automatically recognize locally collinear blocks among organelle genomes and excavate phylogenetically informative regions to construct multiple sequence alignment in a few hours. In addition, HomBlocks supports organelle genomes without annotation and makes adjustment to different taxon datasets, thereby enabling the inclusion of as many common genes as possible. Topology comparison of trees built by conventional multi-gene and HomBlocks alignments implemented in different taxon categories shows that the same efficiency can be achieved by HomBlocks as when using the traditional method. The availability of Homblocks makes organelle phylogenetic analyses more accessible to non-bioinformaticians, thereby promising to lead to a better understanding of phylogenic relationships at an organelle genome level.

基于己糖激酶与葡萄糖-6-磷酸脱氢酶共表达的辅酶NADPH高效再生

【目的】构建己糖激酶与葡萄糖-6-磷酸脱氢酶的大肠杆菌共表达体系,以葡萄糖为底物实现辅酶NADPH的高效再生。【方法】通过分子生物学方法,克隆己糖激酶HKgs、HKpp基因,并于Escherichia coli BL21(DE3)中表达,再将己糖激酶HKgs、HKpp分别与葡萄糖-6-磷酸脱氢酶Gpd PP共表达,实现NADPH的原位再生。比较两个共表达工程菌的辅酶再生效果,并针对催化活力较高的工程菌BL21(HKgs+Gpd PP)进行表达条件优化。【结果】NADPH再生活力达到856 U/L。该辅酶再生体系与醇脱氢酶Adh R联合催化,使不对称还原4-氯乙酰乙酸乙酯的催化活力提高至原始值的2.5倍。【结论】通过己糖激酶与葡萄糖-6-磷酸脱氢酶在大肠杆菌中的共表达,构建了一个新的NADPH高效再生体系,并用于醇脱氢酶催化的不对称还原反应。     

Optimal computation with attractor networks

We investigate the ability of multi-dimensional attractor networks to perform reliable computations with noisy population codes. We show that such networks can perform computations as reliably as possible--meaning they can reach the Cramér-Rao bound--so long as the noise is small enough. "Small enough" depends on the properties of the noise, especially its correlational structure. For many correlational structures, noise in the range of what is observed in the cortex is sufficiently small that biologically plausible networks can compute optimally. We demonstrate that this result applies to computations that involve cues of varying reliability, such as the position of an object on the retina in bright versus dim light.