优良籼型恢复系转基因植株的获得及其遗传稳定性

报道了用基因枪转化法将雪花莲凝集素基因(gna)转移到优良籼型杂交稻恢复系蜀恢527中．PCR、PCR Southern blotting和Southern blotting等分子检测证明，外源基因以多拷贝单位点方式整合到受体基因组中并稳定遗传到转基因第三代(T2)．转基因第一代植株(TO)在株高和结实率上与相应的组培、种子实生苗植株相比，发生明显的不可遗传的变异，随着繁殖代数的增加，转基因植株恢复到与对照植株一致．Western blotting表明，目的基因在转基因植株中正确表达．蛋白活性分析显示，外源基因在转基因植株中合成的蛋白具有凝血生物活性，含量占可溶性总蛋白的0．1％左右．本文还对转基因技术在杂交稻育种中的应用进行了讨沦．图4表2参18     

异源表达胡杨PeCPD基因提高烟草对盐、高氮和干旱胁迫的抗性

油菜素类固醇(Brassinosteroids,BRs)在调节植物生长发育和提高植物对非生物胁迫的抗性中起着重要作用,CPD(Constitutive Photomorphogenesis and Dwarfism)基因编码一种甾醇类23a羟化酶,在BR的生物合成中发挥着重要作用.本研究克隆胡杨Pe CPD基因的c DNA序列,并利用农杆菌介导法,将Pe CPD基因植物过表达载体p BI121-35S::Pe CPD转化野生型烟草,筛选得到Pe CPD基因过表达的转基因烟草植株,并利用Na Cl、高氮和甘露醇分别对转基因和野生型烟草进行胁迫处理,检测转基因烟草的耐受性和相关的生理生化指标.结果表明:(1)在正常条件下,转基因烟草的鲜重是野生型烟草的1.47倍;(2)在Na Cl、高氮以及甘露醇的胁迫处理下,转基因烟草植株的根长、叶片叶绿素和类胡萝卜素的含量均显著高于野生型植株,抗氧化酶SOD和POD活性以及渗透调节物质脯氨酸、可溶性蛋白和可溶性糖的含量也同样表现为转基因烟草显著高于野生型,而野生型植株的丙二醛(MDA)含量显著高于转基因植株.因此在烟草中异源过表达胡杨Pe CPD基因不但能够增加烟草的生物量,而且能够提高烟草对盐、高氮和干旱胁迫的抵抗能力.     

农杆菌介导将高赖氨酸蛋白基因导入谷秆两用水稻

采用农杆菌介导法将高赖氨酸蛋白基因导入到谷秆两用水稻中，GUS组织化学染色、PCR扩增、Southem blot分析表明，该基因已经整合到水稻基因组中，测定9株转基因水稻叶片赖氨酸含量，大部分植株有明显的提高，最高幅度达到了22．71％，图6参15     

瞬时表达比较马铃薯X病毒CP基因3种结构对RNA沉默的诱导效果

RNA沉默是植物抵御病毒侵染的一种防卫机制,病原来源的抗性(pathogen-derived resistance,PDR)被认为是通过RNA沉默起作用的.转化的核酸片段在基因组中的位置、长短和不同排列结构等均影响RNA沉默的效率.用全长马铃薯X病毒(PVX)CP基因构建非翻译的正义、反义和反向重复结构转基因的植物表达载体,通过农杆菌渗入法瞬时表达测试了这些转基因对RNA介导抗性的诱导效率和有效性.结果表明,反向重复的PVXCP是更为有效的RNA沉默诱导因子,同时也证明让目的基因在受试植物中瞬时表达,在转化植物之前能比较快速地检测转基因的表达情况以及表达产物的功能,从而节约时间和资源,并减少盲目性.图3表2参21     

异源表达胡杨PeBAK1;1基因增强烟草对Pst DC3000的抗性

BAK1(BRI1-associated receptor kinase 1)是一个富亮氨酸重复序列(LRR)的膜受体蛋白激酶,除参与植物油菜素内酯信号的转导外,还可以结合其他的LRR-RLKs蛋白来启动植物的先天免疫反应.为明确胡杨Pe BAK1;1基因在烟草中抗Pst DC3000(丁香假单胞杆菌种番茄致病变种)的功能及其在植物抗病中的调控方式,克隆胡杨Pe BAK1;1基因c DNA序列构建Pe BAK1;1基因的过量表达载体p BI121-35S::Pe BAK1;1,并利用农杆菌介导法将其转化野生型烟草,筛选获得Pe BAK1;1基因过表达的转基因烟草植株.生物信息学分析结果表明胡杨Pe BAK1;1蛋白具有植物SERK家族的全部结构特征,系统进化树分析表明胡杨Pe BAK1;1与毛果杨Pt BAK1的序列同源性最高;组织表达分析表明Pe BAK1;1基因主要在根和叶中表达;Pst DC3000侵染试验结果显示,野生型烟草植株接种Pst DC3000后表现出明显的感病症状,而Pe BAK1;1基因过表达的转基因烟草植株表现出明显的抗病症状;Real-time PCR及quantitative real-time-PCR分析结果表明,与野生型相比,转基因烟草中抗病防御相关基因PR1、PR3、PR4和PR5的表达量显著升高,且参与植物生长发育与先天防卫反应的基因BIR1和BON1的表达量也明显升高.上述结果表明,异源表达胡杨Pe BAK1;1基因在烟草抗Pst DC3000过程中起正调控作用,能够增强植物的抗病能力.     

拟南芥VTE1过量表达可以增加维生素E含量和提高烟草植株耐盐行

维生素E在动物细胞内具有抗氧化等重要作用，但在植物体内的功能却鲜为人知．本实验利用CaMV 35S启动子与来源于拟南芥的编码生育酚环化酶（TC）的cDNA（VTE1）构建的嵌合表达载体，以根癌农杆菌介导的叶盘法转化烟草W38．实验结果表明，具有卡那霉素抗性的再生植株经RT-PCR检测，得到了与阳性对照一致的495bp的目标片段；转基因植株的VE含量比对照植株高2倍左右，个别株系高达11倍．实验还发现，在耐盐性实验中转基因植株对盐的抗性明显高于野生型烟草；同时，在不同盐浓度（150、250mmoL／L）胁迫下转基因植株VE含量比未转化植株增加了1．3～1．8倍，首次证明VTE1与植物耐盐性之间的关系．     

水稻锌指蛋白基因OsWIP6的克隆、表达及RNAi载体转化

为了解水稻C2H2型锌指蛋白转录因子基因的表达调控机理及生物学功能,克隆了水稻的一个C2H2型锌指蛋白转录因子基因(OsWIP6),并构建其RNA干涉植物表达载体,通过农杆菌介导转化获得转基因植株.生物信息学分析显示,OsWIP6氨基酸序列与拟南芥两个WIP家族转录因子高度同源.组织差异性表达分析表明,该基因在水稻花发育上具有表达偏向性.与野生型相比,转基因水稻表现为抽穗延迟,结实率及千粒重降低,半定量分析显示转基因植株OsWIP6表达量明显下降.因此,推断OsWIP6基因与水稻育性密切相关.     

高赖氨酸蛋白基因在转基因生菜中的表达和遗传转化

以生菜(LactucasativaL.)为受体材料,将含有高赖氨酸蛋白基因的植物表达载体pLBI用农杆菌介导法进行遗传转化,获得47株转基因生菜植株.PCR和Southern-blotting检测证实目的片段在T0代的整合,RT-PCR检测表明目的基因的转录.T0代植株赖氨酸(Lys)含量的测定分析表明,赖氨酸有不同程度的提高,比对照提高最高的可达9.46%.T1代植株PCR检测证明,目的基因能够遗传.图5表1参25     

番茄SIZ1-like1基因的克隆与功能

SUMO(Small ubiquitin-related modifi er)化修饰是植物体内一种重要的蛋白质功能调节方式.它调控植物细胞中蛋白的降解与定位,植物抗性及激素调节等.SIZ1是SUMO的E3连接酶并在SUMO化中起重要作用.为了解SIZ1基因在番茄中的功能,成功克隆番茄(Solanum lycopersicum)SIZ1基因(SIZ1-like1)并构建番茄SIZ1-like1RNA干涉和过表达载体后,通过农杆菌介导转入野生型番茄,成功获得6个独立的转基因阳性植株.荧光定量PCR(Real-time PCR)分析野生型番茄中SIZ1-like1基因的组织表达特异性,发现SIZ1-like1在植物的各个组织中都有表达.构建SIZ1-like1黄色荧光蛋白融合表达载体,通过对SIZ1-like1融合黄色荧光蛋白转基因番茄的根尖进行荧光显微观察,证实番茄SIZ1蛋白定位于细胞核.干旱胁迫实验分析显示,过表达转基因植株抗旱性强于野生型,且脯氨酸含量是野生型的3倍左右,而RNAi植株抗旱能力则较弱.因此SIZ1基因对番茄抗旱起到了正调控作用.     

BnGID1基因过量表达对甘蓝型油菜矮化突变体NDF-1株高的恢复

在正常株高油菜中克隆了与拟南芥矮化突变相关基因AtGID1同源的BnGID1基因全长cDNA目的片段,将其正向插入表达载体pFGC5941上的CaMV35S启动子下游,构成该基因过量表达载体pFGC5941-BnGID1.经根癌农杆菌介导,导入甘蓝型油菜矮化突变体NDF-1基因组,通过PCR及半定量RT-PCR检测,确定得到抗性转基因植株23株.移至大田生长,观察结果显示,转基因植株明显高于对照矮化突变体NDF-1,平均高出49.05cm.结果表明,BnGID1基因过量表达能够使甘蓝型油菜矮化突变体NDF-1株高得到较大程度的恢复.图7表2参16     

Achieving Open Access to Conservation Science

Conservation science is a crisis discipline in which the results of scientific enquiry must be made available quickly to those implementing management. We assessed the extent to which scientific research published since the year 2000 in 20 conservation science journals is publicly available. Of the 19,207 papers published, 1,667 (8.68%) are freely downloadable from an official repository. Moreover, only 938 papers (4.88%) meet the standard definition of open access in which material can be freely reused providing attribution to the authors is given. This compares poorly with a comparable set of 20 evolutionary biology journals, where 31.93% of papers are freely downloadable and 7.49% are open access. Seventeen of the 20 conservation journals offer an open access option, but fewer than 5% of the papers are available through open access. The cost of accessing the full body of conservation science runs into tens of thousands of dollars per year for institutional subscribers, and many conservation practitioners cannot access pay‐per‐view science through their workplace. However, important initiatives such as Research4Life are making science available to organizations in developing countries. We urge authors of conservation science to pay for open access on a per‐article basis or to choose publication in open access journals, taking care to ensure the license allows reuse for any purpose providing attribution is given. Currently, it would cost $51 million to make all conservation science published since 2000 freely available by paying the open access fees currently levied to authors. Publishers of conservation journals might consider more cost effective models for open access and conservation‐oriented organizations running journals could consider a broader range of options for open access to nonmembers such as sponsorship of open access via membership fees. Obtención de Acceso Abierto a la Ciencia de la Conservación     

Representation of the world's biophysical conditions by the global protected area network

Protected areas (PAs) are often implemented without consideration of already existing PAs, which is likely to cause an overrepresentation of certain biophysical conditions. We assessed the representativeness of the current PA network with regard to the world's biophysical conditions to highlight which conditions are underprotected and where these conditions are located. We overlaid terrestrial and marine PAs with information on biophysical conditions (e.g., temperature, precipitation, and elevation) and then quantified the percentage of area covered by the PA network. For 1 variable at a time in the terrestrial realm, high temperature, low precipitation, and medium and very high elevation were underrepresented. For the marine realm, low and medium sea surface temperature (SST), medium and high sea surface salinity (SSS), and the deep sea were underrepresented. Overall, protection was evenly distributed for elevation across the terrestrial realm and SST across the marine realm. For 2 variables at a time, cold and very dry terrestrial environments had mostly low protection, which was also the case for low SST and low and medium SSS across most depths for marine environments. Low protection occurred mostly in the Sahara and the Arabian Peninsula for the terrestrial realm and along the Tropic of Capricorn and toward the poles for the marine realm. Although biodiversity measures are of prime importance for the design of PA networks, highlighting biophysical gaps in current PAs adds a frequently overlooked perspective. These gaps may weaken the potential of PAs to conserve biodiversity. Thus, our results may provide useful insights for researchers, practitioners, and policy makers to establish a more comprehensive global PA network.     

Quantitative composition of the defensive secretion ofBledius species (Coleoptera: Staphylinidae: Oxytelinae) is adapted to naturally occurring predators

Summary The adaptation of defensive secretions to their target organisms was examined for the abdominal gland secretions ofBledius furcatus, B. spectabilis andB. arenarius. Therefore the target organisms of the secretion of theseBledius species (i.e. their predators) had to be identified. At the collection sites examined these were the earwigLabidura riparia, the antCataglyphis bicolor, the flyLispe candicans, different carabids of the generaPogonus, Dichirotrichus, Dyschirius, Bembidion andCalathus and the wading birdsHaematopus ostralegus andCalidris alba. The secretion of the abdominal glands contains the toxin ptoluquinone dissolved in either-dodecalactone and 1-undecene (B. furcatus andB. spectabilis) or in octanoic acid and octyloctanoate (B. arenarius). The ratio of these solvents is species-specific. Application experiments using some of the natural insect predators (L. riparia, C. bicolor, Pogonus, Di. gustavii, Dyschirius) revealed that these solvent ratios provided a more effective deterrent than other possible ratios. Thus by combining the solvents in certain ratios, the capability of cuticular penetration and therefore the effectiveness of the defensive secretions are adapted to their natural targets.     

Democracy versus development: New urban debates

The politics of development includes subtexts of choice, as underpinned by community consensus (or lack of it), against the larger backdrop of a liberal democracy, with its intricate power structures that influence the choice of concepts and often lead to complicity rather than consent. A dilemma exists between the understanding of field-level practitioners and that of academics, whose understandings of progress differ. Progress can (and often does) imply long-term change across generations whereas development is often more a matter of crisis management and delivering tangible results to end-users. This makes it a bone of contention across political systems, irrespective of local or global dynamics. Development can emerge as a tool to be wielded for power and further political mileage at the expense of progress. Progress cannot be studied as a counterpoint to development as the sense of history is often missing in development interventions. Development tends to be piecemeal as its very logic often stems from the rationale of power. This is not to suggest, however, that development is only about politics; it is definitely more than the politics of everyday life and involves all of civil society that requires its critical and immediate attention. In this sense, it has become a priority for politicians with an underlying agenda designed to out-manouevre dissent and all statements of protest.     

A framework to measure the wildness of managed large vertebrate populations

As landscapes continue to fall under human influence through habitat loss and fragmentation, fencing is increasingly being used to mitigate anthropogenic threats and enhance the commercial value of wildlife. Subsequent intensification of management potentially erodes wildness by disembodying populations from landscape-level processes, thereby disconnecting species from natural selection. Tools are needed to measure the degree to which populations of large vertebrate species in formally protected and privately owned wildlife areas are self-sustaining and free to adapt. We devised a framework to measure such wildness based on 6 attributes relating to the evolutionary and ecological dynamics of vertebrates (space, disease and parasite resistance, exposure to predation, exposure to limitations and fluctuations of food and water supply, and reproduction). For each attribute, we set empirical, species-specific thresholds between 5 wildness states based on quantifiable management interventions. We analysed data from 205 private wildlife properties with management objectives spanning ecotourism to consumptive utilization to test the framework on 6 herbivore species representing a range of conservation statuses and commercial values. Wildness scores among species differed significantly, and the proportion of populations identified as wild ranged from 12% to 84%, which indicates the tool detected site-scale differences both among populations of different species and populations of the same species under different management regimes. By quantifying wildness, this framework provides practitioners with standardized measurement units that link biodiversity with the sustainable use of wildlife. Applications include informing species management plans at local scales; standardizing the inclusion of managed populations in red-list assessments; and providing a platform for certification and regulation of wildlife-based economies. Applying this framework may help embed wildness as a normative value in policy and mitigate the shifting baseline of what it means to truly conserve a species.     

Some polyphagous Homoptera gain protection from their natural enemies by feeding on the toxic plantsSpartium junceum andErythrina corallodendrum (Leguminosae)

Summary Interactions were studied among alkaloid-containing legumes (Erythrina corallodendrum andSpartium junceum) and non-toxic plants (Citrus sinensis, Cucurbita moschata andEuphorbia tirucalli), several polyphagous homopterans,Aphis craccivora (Aphididae),Icerya purchasi, I. aegyptiaca (Margarodidae),Lepidosaphes ulmi (Diaspididae) andPlanococcus citri (Pseudococcidae), and some major natural enemies of these homopterans. Significant reductions in survival due to negative effects of alkaloid containing as compared with non-alkaloidal plants were recorded for the predatorsRodolia cardinalis andChilocorus bipustulatus, but not forCryptolaemus montrouzieri (Coleoptera: Coccinellidae),Chrysoperla carnea (Neuroptera: Chrysopidae) andSympherobius sanctus (Neuroptera: Sympherobiidae). The development time of the larvae or pupae ofR. cardinalis, C. carnea andS. sanctus was longer on the toxic plants than on the non-toxic ones. The percentage of parasitism ofA. craccivora collected from the non-alkaloidal plantsVicia palaestina andMelilotus albus was much higher than that onS. junceum. The parasitoid complexes ofA. craccivora differed between both plant groups. The nutritive value of honeydew ofI. purchasi andA. craccivora, as expressed by the life span ofEncyrtus infelix (Hymenoptera: Encyrtidae) adults, was also investigated. Life spans were significantly longer when the wasps fed on honeydew produced on non-alkaloidal plants (C. sinensis andPittosporum tobira) than on alkaloid containing plants whenI. purchasi — but notA. craccivora — was the producer. It is suggested that the chemical defense ofE. corallodendrum andS. junceum is exploited by polyphagous phytophages to reduce predation. In nature, population growth and density of four of the investigated homopterans are conspicuously high when they developed on the alkaloid containing plant species, and very low on non-alkaloid plants. The efficiency of their natural enemies may be reduced by sequestration of alkaloids (or other toxic plant compounds) or their transfer into excreted honeydew. Therefore it is assumed that a generalist phytophagous homopteran may be protected from its natural enemies, although at different rates of efficiency, if it can safely sequester the host allelochemical when it develops on toxic species within its host range.     

Sexual Predators,Energy Development,and Conservation in Greater Yellowstone

Abstract: In the United States, as elsewhere, a growing debate pits national energy policy and homeland security against biological conservation. In rural communities the extraction of fossil fuels is often encouraged because of the employment opportunities it offers, although the concomitant itinerant workforce is often associated with increased wildlife poaching. We explored possible positive and negative factors associated with energy extraction in the Greater Yellowstone Ecosystem (GYE), an area known for its national parks, intact biological diversity, and some of the New World's longest terrestrial migrations. Specifically, we asked whether counties with different economies—recreation (ski), agrarian (ranching or farming), and energy extractive (petroleum)—differed in healthcare (gauged by the abundance of hospital beds) and in the frequency of sexual predators. The absolute and relative frequency of registered sex offenders grew approximately two to three times faster in areas reliant on energy extraction. Healthcare among counties did not differ. The strong conflation of community dishevel, as reflected by in‐migrant sexual predators, and ecological decay in Greater Yellowstone is consistent with patterns seen in similar systems from Ecuador to northern Canada, where social and environmental disarray exist around energy boomtowns. In our case, that groups (albeit with different aims) mobilized campaigns to help maintain the quality of rural livelihoods by protecting open space is a positive sign that conservation can matter, especially in the face of rampant and poorly executed energy extraction projects. Our findings further suggest that the public and industry need stronger regulatory action to instill greater vigilance when and where social factors and land conversion impact biological systems.     

Comparison of the insecticidal effects of water extracted and intact aromatic plants onAcanthoscelides obtectus,a bruchid beetle pest of kidney beans

Summary Acanthoscelides obtectus (Say) is one of the most damaging pests of kidney beans,Phaseolus vulgaris L. worldwide. However, aromatic plants from the families Lamiaceae, Lauraceae, Myrtaceae and Poaceae can protect kidney beans by direct or delayed insecticidal effect, through increased adult mortality and inhibition of beetle reproduction (both oviposition and adult emergence). The efficiency of hydrodistillated and intact plants fromThymus vulgaris andT. serpyllum, Mentha piperata, Rosmarinus officinalis, Satureia hortensis, Eucalyptus globulus, Laurus nobilis, Origanum vulgare, andCymbopogon nardus was compared. For both extracts,Origanum vulgare had the best effect. The insecticidal effect was induced by more than the essential oils because no significant difference was noticed between distilled and intact plants extract. Inhibition of reproduction was particularly important. These results suggest that lipidic as well as non lipidic, allelochemicals, such as phenolics, or non-protein amino-acids, or flavonoids may be involved in the toxicity of aromatic plants to this beetle.     

Rapid,recurring, structured survey versus bioblitz for generating biodiversity data and analysis with a multispecies abundance model

A bioblitz inexpensively and quickly generates biodiversity data, but bioblitzes are often conducted with haphazard, unreplicated sampling. Results tend to be taxonomically, geographically, or temporally biased, lack metadata, and consist of lists of observed taxa that do not enable further analyses or correction for imperfect detection. A rapid, recurring, structured survey (RRSS) uses a structured sampling design and temporal and spatial replication to survey randomly selected sites on a conservation property. We participated in a loosely structured bioblitz and a subsequent RRSS at Big Canoe Creek Nature Preserve in Springville (St. Clair County), Alabama (USA) to compare observed richness derived from the 2 survey approaches. The RRSS data structure enabled us to fit models that accounted for imperfect detection to estimate abundances, occupancy probabilities, and habitat associations. The loosely structured bioblitz data could not be used in such models. We present a new integrated multispecies abundance model that we applied to avian RRSS data. Our model extension enables estimation for the community, employs data augmentation to estimate the number of undetected species, and incorporates covariates. The RRSS generated a more comprehensive and less biased list of observed taxonomic richness than the loosely structured bioblitz (e.g., 73 vs. 45 bird species and 104 vs. 63 insect families from the RRSS vs. loosely structured bioblitz, respectively). Models fit to the RRSS data identified seasonal patterns in avian community composition and allowed for estimation of habitat–occupancy relationships for insect taxa. The RRSS protocol has potential for broad transferability as a standardized, quick, and inexpensive way to inventory biodiversity and estimate ecological parameters while providing an outreach opportunity.     

Sequestration of cucurbitacin analogs by New and Old World chrysomelid leaf beetles in the tribe Luperini

Summary Two South American polyphagous leaf beetles,Diabrotica speciosa andCerotoma arcuata, selectively accumulated the bitter tasting compound 23,24-dihydrocucurbitacin D in their body after ingesting root tissues of cucurbit plants. Similarly, three Asian Cucurbitaceae-feeding specialists in the genusAulacophora were found to sequester the same compound. Cucurbitacin analogs were shown to deter feeding by a bird predator, indicating an allomonal role for these compounds in cucurbitacin-associated chrysomelid leaf beetles both of New and Old Worlds. The strong affinity to cucurbitacins, selective sequestration of the analogs and consequent protection from predators suggested an ecological adaptation mechanism developed in common among these two geographically isolated subtribes in the Luperini.