首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 78 毫秒
1.
从嗜热子囊菌光孢变种(Thermoascus aurantiacus var.levisporus)RNA中通过RT-PCR克隆出β-葡萄糖苷酶基因bgl Ⅰ的全长序列,cDNA序列为2 672 bp.Genbank登录号为EU269025,将该片段插入巴斯德毕赤酵母Pichia pastoris分泌型表达载体pPIC9K中,获得重组质粒pPIC9K/bgl,经线性化后用电穿孔法导入毕赤酵母GS115中,在醇氧化酶AOXI基因启动子作用下,获得高效表达β-葡萄糖苷酶的毕赤酵母工程菌株.经DEAE-Sepharose Fast Flow阴离子层析纯化了该重组表达蛋白.SDS-PAGER测得该重组蛋白相对分子质量(M)约为120×103.经甲醇诱导,培养基中β-葡萄糖苷酶的活力可达1.2 U/mg,小规模发酵量达0.45 mg/mL.该酶的最适反应温度为60℃,最适反应pH为5.0.于70℃保温30 min仍保持80%的酶活力,具有较高的热稳定性,在pH 3.0~9.0的条件下酸碱耐受性强.图6表1参22  相似文献   

2.
大鼠血红素加氧酶-1基因在乳酸乳球菌中的克隆与表达   总被引:2,自引:0,他引:2  
采用RTPCR技术从大鼠脾总RNA中分离扩增HO1基因,将该基因克隆进pGEMTeasy质粒中,转化大肠杆菌DH5α,提取质粒,分析基因,酶切后与含有乳酸乳球菌启动子NisA的pSEC质粒连接,经电击转化,将重组质粒转入乳酸乳球菌NZ9000中,转化子在含有氯霉素的脑心浸液培养基上培养.用Nisin诱导HO1表达,SDSPAGE和Westernblot分析、鉴定表达产物,并且用分光光度法测定工程菌表达的HO1活性为0.45nmolmg(protein)-1h-1.图5参18  相似文献   

3.
利用PCR技术得到嗜热脂肪芽孢杆菌 (Bacillusstearothermophilus)过氧化氢酶基因 perA ,将该基因与表达载体 pKK2 2 3 3连接构建重组质粒pK perA ,转化大肠杆菌过氧化氢酶HPⅠ和HPⅡ双缺突变株UM 2 ,得到重组大肠杆菌UM 2 1.酶活测定结果表明 ,表达产物具有正常的生物学活性 .SDS PAGE电泳结果显示出明显的特异性表达条带 ,单体Mr =86× 10 3 ,与嗜热脂肪芽孢杆菌所产酶相同 .实验表明 ,重组质粒在宿主UM 2中有较好的稳定性 ,在无选择压力条件下传代 6 0次基本保持稳定 ,传代 10 0次重组质粒保留 80 %以上 .摇瓶实验确定重组菌的最佳表达条件为 :IPTG浓度 ,0 .75mmol/L ;诱导时间 3h ;培养基起始 pH 6 .5 ;诱导温度 37℃ ;装液量 5 0mL/ 2 5 0mL .在优化条件下 ,重组菌产生的过氧化氢酶占菌体总蛋白的 8% ,酶活力可达 35U/mL ,是原始菌株BacillusstearothermophilusIAM110 0 1的 11.7倍 .图 2表 1参 10  相似文献   

4.
通过单因子和多因子摇瓶正交优化试验,确定了米曲霉液态发酵产氨基酰化酶的最佳发酵条件.优化发酵培养基组成(ρ/gL-1):葡萄糖40,蔗糖10,可溶性淀粉20,蛋白胨2.5,马铃薯液1000mL,pH自然.培养基装量50mL/250mL三角瓶,接种量4%.培养温度30℃,转速100r/min,发酵时间42h.每50mL培养物的总酶活由优化前的2627u提高到7338u,是优化前的2.79倍.研究了米曲霉氨基酰化酶的部分酶学性质.该酶催化反应的最适pH为7.0,最适温度为40℃,低浓度的Co2 (5×10-4mol/L)对酶活激活作用显著.图5表2参8  相似文献   

5.
β-甘露聚糖酶基因在枯草芽孢杆菌中的克隆及表达   总被引:2,自引:0,他引:2  
从Bacillus subtilis JNA 3-10中克隆出β-甘露聚糖酶基因成熟肽链编码序列manA1和含信号肽的β-甘露聚糖酶基因manA2,在B.subtilis 168中克隆表达,分别筛选获得高效分泌表达β-甘露聚糖酶的重组菌株BPM1001(pMA5-manA1/B.subtilis 168)和BPM1002(pMA5-manA2/B.subtilis 168),结果表明菌株BPM1002总酶活力是菌株BPM1001的9.65倍,是原始菌株的13.1倍.在基因manA2下游引入His序列克隆出β-甘露聚糖酶基因manA3,获得枯草芽孢杆菌168重组菌株BPM1003.采用Ni-NTA柱纯化重组菌株BPM1003分泌表达的β-甘露聚糖酶,并研究其酶学性质,该酶促反应的最适pH为6.5,最适温度为65℃,在37℃条件下保存一个月酶活力依然保留有77.8%.5 L发酵罐放大实验结果表明魔芋粉对于产β-甘露聚糖酶具有明显的诱导作用,酶活力最高可达2 748.82 U/mL.图9表3参19  相似文献   

6.
耐碱性木聚糖酶产生菌的筛选及发酵条件研究   总被引:5,自引:1,他引:4  
利用刚果红透明圈法,从造纸厂碱性土壤中筛选到一株木聚糖酶生产菌株X24-14,经培养特性研究及16S rDNA序列分析,初步认为该菌属于纤维菌属(Cellulosimicrobium).经发酵条件优化,该菌在麸皮60 g/L,蛋白胨10 g/L,K2HPO4 7.0 g/L, pH 8.5,接种量为5%, 37 ℃, 200 r/min的条件下发酵培养108 h,可达到最大活力,为2 204 u/mL. 该酶最适反应温度为60 ℃;具有较宽的pH作用范围,在pH 4.2~9.4范围内能保持较高的酶活力,在pH 9.4条件下,仍具有80%的酶活力; pH稳定性较好,在4 ℃、pH 11.0的条件下处理24 h仍能保持75%的酶活力.图3表1参12  相似文献   

7.
1,3-丙二醇(1,3-PD)是一种重要的化工原料,其最重要的用途是作为合成聚酯PTT的单体.由于微生物发酵法生产1,3-PD具有操作简单,不易产生有毒副产物等特点,已得到广泛关注.本研究在前期工作的基础上,分别获得了来源于肺炎克雷伯氏菌的甘油脱水酶编码基因dhaB和来源于大肠杆菌的1,3-PD氧化还原酶同工酶编码基因yqhD,利用温控表达载体pBV220串联构建了重组质粒pBV220-yqhD-dhaB,将其转化大肠杆菌得到产1,3-丙二醇温控重组大肠杆菌JM109(pBV220-yqhD-dhaB).该重组菌在LB培养基中,30℃好氧培养12 h至对数生长中期,再经42℃好氧诱导发酵4 h,测得胞内甘油脱水酶和1,3-丙二醇氧化还原酶同工酶的酶活力分别达到260 U/mg蛋白和140U/mg蛋白;在含甘油40 g/L的发酵培养基中,30℃好氧培养12 h至对数生长中期,再经42℃好氧诱导发酵4 h,测得发酵液中1,3-PD含量为8.5 g/L.这将为进一步构建基因工程菌生产1,3-PD打下坚实的基础.图6表1参18  相似文献   

8.
琥珀酸是一种重要的化工产品,微生物发酵法生产琥珀酸具有广阔的应用前景.野生大肠杆菌厌氧发酵产物中琥珀酸含量低,且有大量副产物.为构建高产琥珀酸的重组大肠杆菌,阻断代谢旁路,利用Xer/dif重组酶系统对Escherichia coli CICIM B0013进行代谢途径的调控研究.结果显示,敲除了包括乙酸激酶和磷酸乙酰转移酶基因(ackA-pta)、乳酸脱氢酶基因(ldhA)、丙酮酸甲酸裂解酶基因(pflB)、乙醇脱氢酶基因(adhE)等琥珀酸合成竞争途径中关键酶基因的重组大肠杆菌菌株E.coli CICIM B0013-1040,其发酵液中琥珀酸成为主要产物得到积累;进一步对其PTS系统进行修饰并适当增强PEP羧化酶基因(ppc)表达剂量,成功获得一株具备较强琥珀酸生产能力的菌株E.coli CICIM B0013-1050(pTH-ppc),该菌株厌氧转化葡萄糖36 h,琥珀酸产量达到36.2 g/L,生产强度为1.01 g L-1 h-1,葡萄糖-琥珀酸转化率为64.3%,发酵液经高效液相色谱(HPLC)检测,无乳酸、乙酸、甲酸、乙醇生成.  相似文献   

9.
为获得L 山梨糖还原酶 (SR)整个操纵子序列 ,以 pGEM 3zf( )作为载体构建了Gluconobactersp .S6基因文库 .结合转化子在以L 山梨糖为唯一碳源培养基上的生长情况 ,利用PCR技术筛选到一个含L 山梨糖还原酶基因的阳性克隆pGEM 3zf( ) sr3/E .coli,酶切鉴定插入片断长度约 5 .0kb左右 .以 pGEM 3zf( ) sr3/E .coli质粒DNA为模板 ,扩增SR结构基因 .PCR产物经测序验证为SR基因序列后 ,与表达载体Pet32a( )相连 ,转化宿主大肠杆菌AD4 94 (DE3)对SR基因进行表达 ,并利用重金属离子亲和层析技术对SR融合蛋白进行了纯化 ,对其酶学特性进行了研究 .结果表明 :还原型辅酶II(NADPH)是SR酶蛋白的最适电子供体 ;SR酶蛋白的最适反应pH为 7.0 ,pH为6 .5时保持稳定 ,酶活力较高 ;最适反应温度是 5 0℃ ,30℃时热稳定性较好 .SDS PAGE电泳分析 ,SR融合蛋白的Mr 在6 5× 10 3 左右 ,由此推测出天然SR的Mr 在 5 3× 10 3 左右 ,与SR基因序列推测出的分子量大小一致 .图 13表 2参 6  相似文献   

10.
枯草芽孢杆菌Bacillus subtilis RF1是课题组前期通过基因工程改造得到的一株核黄素高产菌,为了进一步提高核黄素的产量,需要对该菌株进行进一步的基因工程改造.本研究首先将编码的链丝菌素乙酰基转移酶基因sat克隆到p MA5质粒上,构建具有诺瓦丝菌素Norseothricin(NTC)抗性的重组质粒p MA5-sat,实验证实该重组质粒能够用于B.subtilis RF1的抗性筛选.随后将核黄素合成相关的关键酶葡萄糖-6-磷酸脱氢酶编码基因zwf克隆到重组质粒p MA5-sat上,获得重组质粒p MA5-sat-zwf,并成功构建重组菌B.subtilis RF1/p MA5-sat-zwf.结果显示,重组菌胞内葡萄糖-6-磷酸脱氢酶活力比原始菌提高了近50倍,说明葡萄糖-6-磷酸脱氢酶在重组菌中成功过量表达;根据发酵特性分析,重组菌B.subtilis RF1/p MA5-sat-zwf最终核黄素产量达到12.01 g/L,比原始菌B.subtilis RF1提高了30.3%.综上,本研究构建的新型抗性质粒能够成功运用于核黄素生产菌枯草芽孢杆菌的基因工程改造.  相似文献   

11.
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  相似文献   

12.
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.  相似文献   

13.
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.  相似文献   

14.
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.  相似文献   

15.
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.  相似文献   

16.
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.  相似文献   

17.
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.  相似文献   

18.
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.  相似文献   

19.
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.  相似文献   

20.
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.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号