嘌呤霉素抗性基因捕获载体的构建及在细胞中的功能验证

目的构建具有嘌呤霉素抗性基因捕获载体,扩大基因捕获载体的应用范围。方法用经改造的捕获载体（gene trapping vector）稳定转染HepG2．2．15肝癌细胞系,经嘌呤霉素筛选,制作单克隆细胞株。用PCR方法验证该载体的在细胞染色体中的整合,ELISA方法证明捕获载体捕获基因后的细胞的功能改变。结果嘌呤霉素抗性基因捕获载体整合在HepG2．2．15肝癌细胞的染色体上,并能影响细胞HBsAg和HBeAg的分泌。结论新构建的嘌呤霉素抗性基因捕获载体能在具有G418抗性的细胞中捕获有意义的目的基因。     

Model systems for optical trapping: the physical basis and biological applications

The micromechanical methods, among which optical trapping and atomic force microscopy have a special place, are widespread currently in biology to study molecular interactions between different biological objects. Optical trapping is reported to be quite applicable to study the mechanical properties of surface structures onto bacterial (pili and flagella) and eukaryotic (filopodia) cells. The review briefly summarizes the physical basis of optical trapping, as well as the principles of calculating the van der Waals, electrostatic, and donor-acceptor forces when two microparticles or a microparticle and a flat surface are used. Three main types of model systems (abiotic, biotic, and mixed) used in trapping experiments are described, and the peculiarities of manipulation with living (bacteria, fungal spores, etc.) and non-spherical objects (e.g., rod-shaped bacteria) are summarized.     

The Sustainability of Wolverine Trapping Mortality in Southern Canada

Range declines, habitat connectivity, and trapping have created conservation concern for wolverines throughout their range in North America. Previous researchers used population models and observed estimates of survival and reproduction to infer that current trapping rates limit population growth, except perhaps in the far north where trapping rates are lower. Assessing the sustainability of trapping requires demographic and abundance data that are expensive to acquire and are therefore usually only achievable for small populations, which makes generalization risky. We surveyed wolverines over a large area of southern British Columbia and Alberta, Canada, used spatial capture-recapture models to estimate density, and calculated trapping kill rates using provincial fur harvest data. Wolverine density averaged 2 wolverines/1,000 km² and was positively related to spring snow cover and negatively related to road density. Observed annual trapping mortality was >8.4%/year. This level of mortality is unlikely to be sustainable except in rare cases where movement rates are high among sub-populations and sizable un-trapped refuges exist. Our results suggest wolverine trapping is not sustainable because our study area was fragmented by human and natural barriers and few large refuges existed. We recommend future wolverine trapping mortality be reduced by ≥50% throughout southern British Columbia and Alberta to promote population recovery. © 2019 The Authors. The Journal of Wildlife Management published by Wiley Periodicals, Inc. on behalf of The Wildlife Society.     

Specific conjugation of DNA binding proteins to DNA templates through thiol-disulfide exchange

The double-stranded oligodeoxyribonucleotides with single internucleotide disulfide linkages were successfully used for covalent trapping of cysteine containing protein. In particular, an efficient conjugation of DNA methyltransferase SsoII to sequence-specific decoys was demonstrated. The obtained results assume that synthetic oligodeoxyribonucleotides bearing a new trapping site can be used as new tools to study and manipulate biological systems.     

Predator Presence and Vegetation Density Affect Capture Rates and Detectability of Litoria aurea Tadpoles: Wide-Ranging Implications for a Common Survey Technique

Trapping is a common sampling technique used to estimate fundamental population metrics of animal species such as abundance, survival and distribution. However, capture success for any trapping method can be heavily influenced by individuals’ behavioural plasticity, which in turn affects the accuracy of any population estimates derived from the data. Funnel trapping is one of the most common methods for sampling aquatic vertebrates, although, apart from fish studies, almost nothing is known about the effects of behavioural plasticity on trapping success. We used a full factorial experiment to investigate the effects that two common environmental parameters (predator presence and vegetation density) have on the trapping success of tadpoles. We estimated that the odds of tadpoles being captured in traps was 4.3 times higher when predators were absent compared to present and 2.1 times higher when vegetation density was high compared to low, using odds ratios based on fitted model means. The odds of tadpoles being detected in traps were also 2.9 times higher in predator-free environments. These results indicate that common environmental factors can trigger behavioural plasticity in tadpoles that biases trapping success. We issue a warning to researchers and surveyors that trapping biases may be commonplace when conducting surveys such as these, and urge caution in interpreting data without consideration of important environmental factors present in the study system. Left unconsidered, trapping biases in capture success have the potential to lead to incorrect interpretations of data sets, and misdirection of limited resources for managing species.     

Potential of mass trapping for long-term pest management and eradication of invasive species

Semiochemical-based pest management programs comprise three major approaches that are being used to provide environmentally friendly control methods of insect pests: mass trapping, "lure and kill," and mating disruption. In this article, we review the potential of mass trapping in long-term pest management as well as in the eradication of invasive species. We discuss similarities and differences between mass trapping and other two main approaches of semiochemical-based pest management programs. We highlight several study cases where mass trapping has been used either in long-term pest management [e.g., codling moth, Cydia pomonella (L.); pink bollworm, Pectinophora gossypiella (Saunders); bark beetles, palm weevils, corn rootworms (Diabrotica spp.); and fruit flies] or in eradication of invasive species [e.g., gypsy moth, Lymantria dispar (L.); and boll weevil, Anthonomus grandis grandis Boheman). We list the critical issues that affect the efficacy of mass trapping and compare these with previously published models developed to investigate mass trapping efficacy in pest control. We conclude that mass trapping has good potential to suppress or eradicate low-density, isolated pest populations; however, its full potential in pest management has not been adequately realized and therefore encourages further research and development of this technology.     

High throughput gene trapping and postinsertional modifications of gene trap alleles

Gene trapping is a high-throughput insertional mutagenesis approach that has been primarily used in mouse embryonic stem cells (ESCs). As a high throughput technology, gene trapping helped to generate tenth of thousands of ESC lines harboring mutations in single genes that can be used for making knock-out mice. Ongoing international efforts operating under the umbrella of the International Knockout Mouse Consortium (IKMC; www.knockoutmouse.org) aim to generate conditional alleles for every protein coding gene in the mouse genome by high throughput conditional gene targeting and trapping. Here, we provide protocols for gene trapping in ESCs that can be easily adapted to any other mammalian cell. We further provide protocols for handling and verifying conditional gene trap alleles in ESC lines obtained from the IKMC repositories and describe a highly efficient method for the postinsertional modification of gene trap alleles. More specifically, we describe a protein tagging strategy based on recombinase mediated cassette exchange (RMCE) that enables protein localization and protein-protein interaction studies under physiological conditions.     

Electrodeless dielectrophoresis of single- and double-stranded DNA

Dielectrophoretic trapping of molecules is typically carried out using metal electrodes to provide high field gradients. In this paper we demonstrate dielectrophoretic trapping using insulating constrictions at far lower frequencies than are feasible with metallic trapping structures because of water electrolysis. We demonstrate that electrodeless dielectrophoresis (EDEP) can be used for concentration and patterning of both single-strand and double-strand DNA. A possible mechanism for DNA polarization in ionic solution is discussed based on the frequency, viscosity, and field dependence of the observed trapping force.     

High-throughput trapping of secretory pathway genes in mouse embryonic stem cells

High-throughput gene trapping is a random approach for inducing insertional mutations across the mouse genome. This approach uses gene trap vectors that simultaneously inactivate and report the expression of the trapped gene at the insertion site, and provide a DNA tag for the rapid identification of the disrupted gene. Gene trapping has been used by both public and private institutions to produce libraries of embryonic stem (ES) cells harboring mutations in single genes. Presently, approximately 66% of the protein coding genes in the mouse genome have been disrupted by gene trap insertions. Among these, however, genes encoding signal peptides or transmembrane domains (secretory genes) are underrepresented because they are not susceptible to conventional trapping methods. Here, we describe a high-throughput gene trapping strategy that effectively targets secretory genes. We used this strategy to assemble a library of ES cells harboring mutations in 716 unique secretory genes, of which 61% were not trapped by conventional trapping, indicating that the two strategies are complementary. The trapped ES cell lines, which can be ordered from the International Gene Trap Consortium (http://www.genetrap.org), are freely available to the scientific community.     

相机陷阱法在潮间带底栖蟹类群落生态学中的应用研究

潮间带大型底栖动物调查是滨海湿地生物多样性监测的重要环节。目前普遍应用的传统侵入性调查方法（挖掘法、手捕法和陷阱诱捕法）存在干扰强、耗时久和劳动密集等问题,无法客观反映活动性较强的蟹类等重要功能类群的群落结构和物种多样性。对陆地野生动物研究中常用的相机陷阱法进行了符合潮间带应用条件的便携式设计,并以红树林蟹类为例,通过与3种传统侵入性调查方法进行对比,探讨相机陷阱法在潮间带底栖蟹类群落生态学研究中的应用。结果表明：（1）相机陷阱法在种群密度测定方面比挖掘法提高2.1倍,比手捕法提高10.3倍,比陷阱诱捕法提高16.3倍;（2）相机陷阱法在调查物种数方面均高于3种传统侵入性调查方法,适用于沙蟹科（Ocypodidae）和方蟹科（Grapsidae）蟹类调查;（3）相机陷阱法对个体大小选择性弱,采样更加全面;（4）相机陷阱法在群落alpha多样性（Shannon-Wiener指数）、beta多样性（样方-多度矩阵总方差Var （Y））和功能多样性（Rao二次熵和功能离散度）调查方面整体不弱于侵入性调查方法。此外,相较于侵入性调查方法,相机陷阱法对沉积物结构破坏小、不干扰蟹类群落结构,在兼顾数据丰富性的同时又具有高效性。然而,由于相机陷阱法基于蟹类底表活动性,无法适用于冬季调查、夜行性蟹类和游泳型蟹类调查。未来,相机陷阱法的发展将与基于深度学习的图像处理技术结合以满足数据处理需求。研究揭示了相机陷阱法在潮间带底栖蟹类群落中的应用优越性与局限性,为潮间带大型底栖动物群落生态学的研究提供技术支撑。     

Hand searching versus pitfall trapping: how to assess biodiversity of ground beetles (Coleoptera: Carabidae) in high altitude equatorial Andes?

The use of ground beetles (Coleoptera: Carabidae) as bioindicators of environmental change depends on the reliability and the effectiveness of the sampling methods. Those that have been tested in the temperate zone and in tropical forests still await experimentation in tropical high-altitude environments. For the first time, pitfall trapping and hand searching have been compared in Ecuadorian páramo above 4000 m a.s.l., in terms of practical effectiveness. The study was performed on six volcanoes and was based on the comparison of 28 sampling sessions (pitfall trapping and hand searching) performed along two different elevational belts [lower superpáramo (LSP) and upper superpáramo (USP)]. Analyses of sampling sessions showed that detected species richness is slightly higher with hand searching than with pitfall trapping, regardless of the elevation. Additionally, hand searching is more time-effective than pitfall trapping. The performance of the sampling method slightly varies when species assemblage composition is analysed in relation to elevational belts. In the LSP, hand searching and pitfall trapping should be simultaneously used to obtain exhaustive inventories of carabid biodiversity, since different species are likely to be collected by each method. In the USP, hand searching and pitfall trapping efficiency is very similar, but hand searching allows to collect a slightly larger number of species. Lastly, the sample-based rarefaction curves showed that four temporal replicates are mandatory to obtain a robust dataset and an exhaustive inventory of the true species richness and species assemblages composition. Our findings suggest a combined use of hand searching and pitfall trapping in the LSP, while both methods can be used alone for surveying carabids in the USP. Furthermore, hand searching is recommended if the aim is to obtain an inventory of species diversity, whereas pitfall trapping seems more convenient for fine grain ecological and comparative studies.     

Differential trappability of small mammals in three habitats of Southeastern Brazil.

We compared the trappability of marked and unmarked individuals in species of marsupials and rodents of three tropical assemblages of small mammals in Brazil. Two studies used trapping grids, one in cerrado and the other in an Atlantic forest reserve, whereas the study in a rural area used transects. In the two studies using trapping grids, marked animals were frequently more trappable than unmarked ones, but in some species this difference was not significant. In the rural area, marked and unmarked animals did not differ significantly. The number of recaptures per resident animal was higher in the two studies using trapping grids than in the rural area where transects were used. Differences in trappability between the three studies might have been caused by differences not only in trapping design (grids vs. transects), but also in the type of trap used, bait, and habitat. Although differential trappability tends to be considered the rule in small mammals, these results suggest that trappability of marked and unmarked animals may be specific for the particular combination of sampling design, field methods, and habitat under study.     

AZI基因被捕获的鉴定实验

目的 确立基因捕获细胞中被捕获的基因名称. 方法 Southern印迹确定合适的限制性内切酶,用质粒拯救（plasmid rescue）获得含有细胞染色体DNA的质粒,测序.结果 本次实验中,被捕获载体整合的基因是AZI基因. 结论质粒拯救方法能确立质粒整合细胞染色体上准确的位置.     

Excited state dynamics in photosystem I: effects of detergent and excitation wavelength.

Femtosecond transient absorption spectroscopy has been used to investigate the energy transfer and trapping processes in both intact membranes and purified detergent-isolated particles from a photosystem II deletion mutant of the cyanobacterium Synechocystis sp. PCC 6803, which contains only the photosystem I reaction center. Processes with similar lifetimes and spectra are observed in both the membrane fragments and the detergent-isolated particles, suggesting little disruption of the core antenna resulting from the detergent treatment. For the detergent-isolated particles, three different excitation wavelengths were used to excite different distributions of pigments in the spectrally heterogeneous core antenna. Only two lifetimes of 2.7-4.3 ps and 24-28 ps, and a nondecaying component are required to describe all the data. The 24-28 ps component is associated with trapping. The trapping process gives rise to a nondecaying spectrum that is due to oxidation of the primary electron donor. The lifetimes and spectra associated with trapping and radical pair formation are independent of excitation wavelength, suggesting that trapping proceeds from an equilibrated excited state. The 2.7-4.3 ps component characterizes the evolution from the initially excited distribution of pigments to the equilibrated excited state distribution. The spectrum associated with the 2.7-4.3 ps component is therefore strongly excitation wavelength dependent. Comparison of the difference spectra associated with the spectrally equilibrated state and the radical pair state suggests that the pigments in the photosystem I core antenna display some degree of excitonic coupling.     

Trapping intermediates in the crystal: ligand binding to myoglobin

Crystal structures of the reactive short-lived species that occur in chemical or binding reactions can be determined using X-ray crystallography via time-resolved or kinetic trapping approaches. Recently, various kinetic trapping methods have been used to determine the structure of intermediates in ligand binding to myoglobin.     

昆虫的生物光电效应与虫害治理应用

农业虫害的治理需要依据为害昆虫的特性提出与环境适宜、生态兼容的技术体系和关键技术。为害昆虫表现了对敏感光源具有个体差异性和群体一贯性的趋光性行为特征,并通过视觉神经信号响应和生理光子能量需求的方式呈现出生物光电效应的作用本质。利用昆虫的这种趋性行为诱导增益特性,一些光电诱导杀虫灯技术以及害虫诱导捕集技术广泛地应用于农业虫害的防治,具有良好的应用前景。     

Mechanism of A23187-induced airway obstruction in the guinea pig

Exposure of conscious guinea pigs to A23187 aerosol produced a concentration-related increase of excised lung gas volumes (ELGV), i.e., postmortem pulmonary gas trapping. Measurements of ELGV were highly correlated with in vivo measurements of dynamic compliance (Cdyn) and total pulmonary resistance (RL) and were used as an indication of in vivo airway obstruction. We pretreated guinea pigs intravenously with the following drugs: atropine; LY163443, a selective LTD4/E4 antagonist; indomethacin; propranolol; and pyrilamine. The guinea pigs were exposed for 8 minutes to the A23187 aerosol, and ELGV measurements were then made. Atropine or pyrilamine prevented the A23187-induced gas trapping. Indomethacin or propranolol tended to potentiate the response and when combined, they potentiated the gas trapping by 80%. LY163443 had no effect alone, but when combined with indomethacin, propranolol, and pyrilamine, inhibited A23187-induced gas trapping by 67%. We conclude that cholinergic and histaminergic mechanisms play major roles in the ionophore-induced pulmonary gas trapping of the guinea pig. With appropriate pretreatment, sulfidopeptide leukotrienes may produce a substantial effect.     

Removal trapping to control Artioposthia triangulata

Removal trapping was used to try and reduce numbers of the predatory planarian Artioposthia triangulata in polythene tunnels on a nursery and a grass field. Ten months of trapping in the tunnels failed to have any impact upon flatworm numbers but weekly catches reduced over a period of 12 weeks' trapping in grassland. The calculated residual population in the grassland (40 ha^-1) was small in comparison with that estimated from formalin sampling (940 × 1288 ha^-1). A separate field study examined the effect of trap density on catch by spacing traps at 2.5, 5, 10 and 20 m intervals. This showed that the numbers of planarians caught was inversely related to the logarithm of trap spacing. It is concluded that removal trapping is too demanding of resources to be a viable control option for this pest in commercial and agricultural situations.     

Estimating tsetse population parameters: application of a mathematical model with density-dependence

A density-dependent model is used to describe the dynamics of an open population of tsetse flies (Diptera: Glossinidae). Immigration (or emigration) takes place when the total population is below (or above) a biologically determined threshold value. The population is also subjected to birth and death rates, as well as to the risk of being trapped (continuously or intermittently). During trapping the population decreases toward a 'low' equilibrium population and when trapping ceases the population starts recovering and increases toward a 'high' equilibrium. The model is fitted using data collected on trapped flies in four experiments. The first one was conducted with 'intermittent trapping' (i.e. several trapping-recovery cycles) on Glossina fuscipes fuscipes Newstead in the Central African Republic (Bangui area). In the other experiments, trapping data on Glossina palpalis palpalis (Robineau-Desvoidy) was collected in 'aggregate' form over several days at a time. Two of these were in Congo-Brazzaville (Bouenza area) and one in the Ivory Coast (Vavoua focus). Estimates are derived for the low and high equilibrium values as well as the trapping rate. The estimated effect of sustained trapping is to reduce the population to low equilibrium values that are 85-87% lower than the levels without trapping. The effects of the natural intrinsic growth and of the migration flows cannot be estimated separately because in the model they are mathematically indistinguishable.