用RACE技术扩增并克隆牛BMP4基因3'端序列

RACE技术是一项扩增基因末端序列的新技术。该研究从牛BMP4基因出发,以牛软骨的RNA为模板,按照不同物种BMP4基因的相似性设计特异引物,运用PCR和RACE技术扩增并获得了特异片段,该片段经PCR、酶切和测序验证,证实所克隆序列为牛BMP4的3′端序列,包含有1170bp组成的开放读码框(ORF),编码389个氨基酸,3′非编码区121bp个核苷酸和poly(A)15。同源性分析结果表明,牛BMP4 cDNA最大开放读码框所推测的氨基酸序列与已知人、小鼠、大鼠、狗、羊和鸡等真核生物BMP4氨基酸序列进行比较,分别有94.5%、93.1%、91.9%、87.4%、94.2%、79%的同源性。这为克隆其他物种的BMP4基因提供了依据,同时牛骨形态发生蛋白的测序为我们更好的理解牛的生骨机理提供帮助。     

丘陵地区地形梯度上植被格局的分异研究概述

植物群落的本质特征之一是群落中的植物和环境之间存在一定的相互关系。湿润的丘陵地区是由水侵蚀而形成的包含各种干扰频率的生境复合体,作为中尺度的地形单位,可以通过侵蚀前线划分为上部坡面和下部坡面两个小尺度的地形单位,而上部坡面可以进一步划分为顶坡、上部边坡、谷头凹地等微地形单元,下部坡面可以进一步划分为下部边坡、麓坡、泛滥性阶地及谷床等微地形单元。上部坡面发育的是气候顶极群落,沿顶坡向谷头凹地,群落发生逐渐、连续的变化,下部坡面发育的为地形群落,其物种组成、结构以及其它生态特征与上部坡面具有显著的差异,而其微地形单元之间植被的变化不明显。干扰作用是不同地形植被分异的控制因子,也是地形植被维持和更新的关键因子。下部坡面以相对积极的土壤侵蚀、滑坡和崩塌等过程为特征,其植被更新依赖于频繁的地面干扰,而上部坡面长期稳定,其植被更新依赖于林窗动态。地形是影响植被格局的最重要的也是最基本的生境因子,其引起的生境生态位分化为物种的共存提供了条件,导致了小尺度空间内高生物多样性的形成和维持。     

基于NOAA-AVHRR数据的中国东部地区植被遥感分类研究

该文采用 19幅 (时间跨 8个月 ) 时间序列的NOAA_AVHRR的归一化植被指数 (NDVI) 最大值合成影像遥感数据, 经过主分量分析 (Principlecomponentanalysis, PCA) 处理后, 用非监督分类方法的ISODATA算法, 对中国东部地区的 (五省一市 ) 植被进行分类, 结果可以分出 2 8种土地覆盖类型, 除了两种类型为水体和城市或裸地外, 其余 2 6种类型均为植被类型, 根据中国植被分类系统, 这 2 6类可以归并为 6大植被类型 :1) 常绿阔叶林 ;2 ) 针叶林 ;3) 竹林 ;4 ) 灌草丛 ;5 ) 水生植被 ;6 ) 农业植被。用 1∶10 0 0 0 0 0数字化《中国植被图集》的植被类型检验遥感分类结果表明, 针叶林、灌草丛、常绿阔叶林和农业植被的分类具有较高的位置精度和面积精度, 位置精度分别为 79.2 %、91.3%、6 8.2 %和 95.9%, 面积精度分别达到 92.1%、95.9%、6 3.8%和 90.5 %。这 6大植被类型在地理空间上的分布规律与中国东部常绿阔叶林区植被的地带性分布基本一致。     

Horizontal transmission, vertical inactivation, and stochastic loss of mariner-like transposable elements

Horizontal transmission has been well documented as a major mechanism for the dissemination of mariner-like elements (MLEs) among species. Less well understood are mechanisms that limit vertical transmission of MLEs resulting in the "spotty" or discontinuous distribution observed in closely related species. In this article we present evidence that the genome of the common ancestor of the melanogaster species subgroup of Drosophila contained an MLE related to the mellifera (honey bee) subfamily. Horizontal transmission, approximately 3-10 MYA, is strongly suggested by the observation that the sequence of the MLE in Drosophila erecta is 97% identical in nucleotide sequence with that of an MLE in the cat flea, Ctenocephalides felis. The D. erecta MLE has a spotty distribution among species in the melanogaster subgroup. The element has a high copy number in D. erecta and D. orena, a moderate copy number in D. teissieri and D. yakuba, and was apparently lost ("stochastic loss") in the lineage leading to D. melanogaster, D. simulans, D. mauritiana, and D. sechellia. In D. erecta, most copies are concentrated in the heterochromatin. Two copies from D. erecta, denoted De12 and De19, were cloned and sequenced, and they appear to be nonfunctional ("vertical inactivation"). It therefore appears that the predominant mode of MLE evolution is vertical inactivation and stochastic loss balanced against occasional reinvasion of lineages by horizontal transmission.      

A phylogenetic approach to the identification of phosphoglucomutase genes

The expanding molecular database provides unparalleled opportunities for characterizing genes and for studying groups of related genes. We use sequences drawn from the database to construct an evolutionary framework for examining the important glycolytic enzyme phosphoglucomutase (PGM). Phosphoglucomutase plays a pivotal role in the synthesis and utilization of glycogen and is present in all organisms. In humans, there are three well-described isozymes, PGMI, PGM2, and PGM3. PGM1 was cloned 5 years ago; however, repeated attempts using both immunological approaches and molecular probes designed from PGM1 have failed to isolate either PGM2 or PGM3. Using a phylogenetic strategy, we first identified 47 highly divergent prokaryotic and eukaryotic PGM-like sequences from the database. Although overall amino acid identity often fell below 20%, the relative order, position, and sequence of three structural motifs, the active site and the magnesium-- and sugar-binding sites, were conserved in all 47 sequences. The phylogenetic history of these sequences was complex and marked by duplications and translocations; two instances of transkingdom horizontal gene transfer were identified. Nonetheless, the sequences fell within six well-defined evolutionary lineages, three of which contained only prokaryotes. Of the two prokaryotic/eukaryotic lineages, one contained bacterial, yeast, slimemold, invertebrate, and vertebrate homologs to human PGM1 and the second contained likely homologs to human PGM2. Indeed, an amino acid sequence, derived from a partial human cDNA, that fell within the second cross-kingdom lineage bears several characteristics expected for PGM2. A third lineage may contain homologs to human PGM3. On a general level, our phylogenetic-based approach shows promise for the further utilization of the extensive molecular database.      

Short chain oligogalacturonides induce ethylene production and expression of the gene encoding aminocyclopropane 1-carboxylic acid oxidase in tomato plants

Oligogalacturonic acids (OGAs), derived from plant cell wall pectin, have been implicated in a number of signal transduction pathways involved in growth, development and defense responses of higher plants. This study investigates the size range of OGAs capable of inducing ethylene synthesis in tomato plants, and demonstrates that in contrast with many other effects, only short chain OGAs are active. Oligomers across a range of DP from 2-15 were separated and purified to homogeneity by QAE-Sephadex anion exchange chromatography using a novel elution system. The OGAs were applied to tomato plants and assayed for their ability to induce ethylene gas release and changes in steady state levels of mRNA encoding the ethylene forming enzyme aminocyclopropane-1-carboxylic acid oxidase (ACO). The study demonstrated that only OGAs in the size range of DP4-6 were active both in eliciting ACO expression and in the production of ethylene.      

Release of Membrane-Bound Trails by Trypanosoma cruzi Amastigotes onto Modified Surfaces and Mammalian Cells

ABSTRACT. Upon incubation at 37° C onto glass coverslips coated with Concanavalin A, poly-L-lysine, or a monoclonal antibody (1D9) directed to the parasite major surface glycoprotein Ssp-4, extracellular Trypanosoma cruzi amastigotes release trails of material barely visible by light microscopy. This release is not associated with parasite movements. Immunolabeling studies confirmed that the material is derived from the parasite's membrane since thin section through samples labeled with 1D9 revealed that the trails are membrane-bound structures. Scanning electron microscopy showed that the ∼0.1-μm thick trails of material emerging from the amastigotes can be uniform or beaded, indicating a tendency to vesiculation. The trails are preferentially released from the flagellar pocket region and/or at the opposite posterior end of the parasite body, and seem to be devoid of microtubules. The release is time and temperature-dependent and fixed parasites do not form trails. All attempts to inhibit trail release using drugs (antimycin A, sodium azide, cytochalasin D, nocodazole, genistein, staurosporine, EGTA) failed. The observation of trails associated with intracellular parasites and amastigotes invading Vero cells suggests that this is probably a physiological process.     

Interaction of ouabain with the Na(+) pump in intact epithelial cells

To determine the specificity and efficacy of [(3)H]ouabain binding as a quantitative measure of the Na(+) pump (Na(+), K(+)-ATPase) and as a marker for the localization of pumps involved in transepithelial Na(+)-transport, we analyzed the interaction of [(3)H]ouabain with its receptor in pig kidney epithelial (LLC-PK(1)) cells. When these epithelial cells are depleted of Na(+) and exposed to 2 muM [(3)H]ouabain in a Na(+)-free medium, binding is reduced by 90 percent. When depleted of K(+) and incubated in a K(+)- free medium, the ouabain binding rate is increase compared with that measured at 5 mM. This increase is only demonstable when Na(+) is present. The increased rate could be attributed to the predominance of the Na(+)-stimulated phosphorylated form of the pump, as K(+) is not readily available to stimulate dephosphorylation. However, some binding in the K(+)-free medium is attributable to pump turnover (and therefore, recycling of K(+)), because analysis of K(+)-washout kinetics demonstrated that addition of 2 muM ouabain to K(+)-depleted cells increased the rate of K(+) loss. These results indicate that in intact epithelial cells, unlike isolated membrane preparations, the most favorable condition for supporting ouabain binding occurs when the Na(+), K(+)-ATPase is operating in the Na(+)-pump mode or is phosphorylated in the presence of Na(+). When LLC-PK(1) cells were exposed to ouabain at 4 degrees C, binding was reduced by 97 percent. Upon rewarming, the rate of binding was greater than that obtained on cells kept at a constant 37 degrees C. However, even at this accelerated rate, the time to reach equilibrium was beyond what is required for cells, swollen by exposure to cold, to recover normal volume. Thus, results from studies that have attempted to use ouabain to eliminate the contribution of the conventional Na(+) pump to volume recovery must be reevaluated if the exposure to ouabain was done in the cold or under conditions in which the Na(+) pump is not operating.     

Fast accumulation of few polyhedra mutants during passage of a Spodoptera frugiperda multicapsid nucleopolyhedrovirus (Baculoviridae) in Sf9 cell cultures

Baculoviruses are a group of viruses that infect invertebrates and that have been used worldwide as a biopesticide against several insect pests of the Order Lepidoptera. In Brazil, the baculovirus Spodoptera frugiperda multicapsid nucleopolyhedrovirus (SfMNPV; Baculoviridae) has been used experimentally to control S. frugiperda (Lepidoptera: Noctuidae), an important insect pest of corn (maize) fields and other crops. Baculoviruses can be produced either in insect larvae or in cell culture bioreactors. A major limitation to the in vitro production of baculoviruses is the rapid generation of mutants when the virus undergoes passages in cell culture. In order to evaluate the potential of in vitro methods of producing SfMNPV on a large-scale, we have multiplied a Brazilian isolate of this virus in cell culture. Extensive formation of few polyhedra mutants was observed after only two passages in Sf9 cells.     

Photosynthesis of oak trees [Quercus petraea (Matt.) Liebl.] during drought under field conditions: diurnal course of net CO2 assimilation and photochemical efficiency of photosystem II

Adult trees of Quercus petraea were submitted to controlled water shortage in a natural stand near Nancy, France. Diurnal course of net CO₂ assimilation rate (A) was measured in situ together with chlorophyll a fluorescence determined on dark adapted leaves. In 1990, trees experienced a strong water stress, with predawn and midday leaf water potentials below –2·0 and –3·0 MPa, respectively. Diurnal course of A of well-watered trees exhibited sometimes important midday decreases in A related to high temperature and vapour pressure deficit. Decreases in initial (F_o) and maximal (F_m) fluorescence and sometimes in photochemical efficiency of photosystem II (F_v/F_m) were observed and probably revealed the onset of mechanisms for thermal de-excitation. These mechanisms were shown to be sensitive to dithiothreitol. All these effects were reversible and vanished almost completely overnight. Therefore, they may be considered as protective mechanisms adjusting activity of photosystem II to the electron requirement for photosynthesis. Water stress amplified these reactions: A was strongly decreased, showing important midday depression; diurnal reductions in F_m and F_v/F_m were enhanced. The same trends were observed during summer 1991, despite a less marked drought. These protective mechanisms seemed very effective, as no photoinhibitory damage to PS II could be detected in either water stressed or control trees.