Fitness Implications of Clonal Integration and Leaf Dynamics in a Stoloniferous Herb, Nelsonia canescens (Lam.) Spreng (Nelsoniaceae)

Very little has been published on the life-history significance of clonal plants inhabiting southern African savanna environments. This study investigated the fitness implications of clonal integration, resprouting behaviour and growth phenology in a stoloniferous herb, Nelsonia canescens (blue pussyleaf) at a savanna site in Zambia, central Africa. Census data on growth and survival were obtained regularly on permanently marked ramets over a 4-year period, from 2001 to 2005, and analyzed to assess how physiological integration and module demography contribute to fitness in Nelsonia. Above ground and below ground growth occurred during the dry and rainy seasons, respectively. Dry season growth was characterized by resprouting and production of stolons that bore small pubescent leaves with high mortality (30–80% month⁻¹). Deep roots and high leaf turnover appear to contribute to sustained growth during the dry season when topsoil moisture and nutrient availability are low. The interaction between maximum temperature and precipitation explained a significant proportion (59%, p<0.01) of the monthly variation in leaf size and increasing evapo-transpiration levels appeared to trigger the shift in leaf size from a large wet season type to a small dry season one. During the dry season Nelsonia resprouted from dormant buds buried at the time of root development in daughter ramets in the rainy season. Temporal integration significantly (p<0.05) enhanced survival of daughter ramets. However, daughter ramets with severed mother–daughter ramet inter-connectors experienced high initial mortality that was caused by both early stolon severing and drought stress during the root development phase. The majority of ramets lived for 5–10 months and 25% were still alive at the age of 3.5 years. The study showed that although the growth phenology of Nelsonia has serious ecological implications for accessing scarce resources during the dry season, the species utilizes a number of strategies to overcome resource limitations in a seasonally heterogenous environment. Co-ordinating editor: G. P. Cheplick     

Effects of resource heterogeneity on nitrogen translocation within clonal fragments of Sasa palmata: an isotopic (15N) assessment

BACKGROUND AND AIMS: Clonal fragments of the rhizomatous dwarf bamboo Sasa palmata, which widely predominates in temperate regions of Japan, were grown under heterogeneous resource conditions such as gap understories or nutrient-patchy grassland. Clonal fragments develop multiple ramets with long rhizomes and appear to be physiologically integrated by the translocation of assimilates. The glasshouse experiment reported here was designed to clarify the mechanisms of physiological integration of nitrogen more precisely. METHODS: To assess how resource conditions influence the amount of nitrogen translocation, and which organ acts as the strongest sink, two experiments were conducted that traced movement of 15N label between interconnected pairs of ramets to compare homogeneous and heterogeneous light and soil nitrogen conditions. KEY RESULTS: The amount of 15N translocated to leaves was between 9% and 11% greater in high-N and high-light ramets in the heterogeneous compared with homogeneous treatments. Under heterogeneous soil nitrogen conditions, translocation increased from individual ramets in resource-rich patches to ramets in resource-poor patches, while the reverse was true under heterogeneous light environments, reflecting differences in the positions of leaves that act as the strongest sinks. Neither the mass increments nor the total mass of clonal fragments was significantly affected by heterogeneity of either light or nutrients, possibly because the experimental period was too short for differences to manifest themselves. CONCLUSIONS: This study clearly demonstrated that nitrogen is readily translocated between ramets, particularly under heterogeneous resource conditions. The translocation patterns were governed by functional 'division of labour' mechanisms that resulted in net nitrogen movement from understory sites to gaps, thereby enhancing the carbon acquisition of the whole fragment. Thus, physiological integration may provide benefits for S. palmata when it is growing under heterogeneous conditions in which there are deficits of certain environmental resources.     

Integration of visual and olfactory cues of hosts and non-hosts by three bark beetles (Coleoptera: Scolytidae)

Abstract.  1. There has been a long-standing pre-occupation with how phytophagous insects use olfactory cues to discriminate hosts from non-hosts. Foragers, however, should use whatever cues are accurate and easily assessed, including visual cues.
2. It was hypothesised that three bark beetles, the mountain pine beetle (MPB), Dendroctonus ponderosae Hopkins, the Douglas-fir beetle (DFB), D. pseudotsugae Hopkins, and the western balsam bark beetle (WBBB), Dryocoetes confusus Swaine, integrate visual and olfactory information to avoid non-host angiosperms (e.g. paper birch, trembling aspen), that differ in visual and semiochemical profile from their respective host conifers (lodgepole pine, Douglas-fir, interior fir), and tested this hypothesis in a series of field trapping experiments.
3. All three species avoided attractant-baited, white (non-host simulating) multiple-funnel traps, and preferred attractant-baited black (host-simulating) traps. In experiments combining white, non-host traps with non-host angiosperm volatiles, bark beetles were repelled by these stimuli in an additive or redundant manner, confirming that these species could integrate visual and olfactory information to avoid non-host angiosperms while flying.
4. When antiaggregation pheromones were released from white traps, the DFB and MPB were repelled in an additive-redundant manner, suggesting that beetles can integrate diverse and potentially anomalous stimuli.
5. The MPB demonstrated the most consistent visual preferences, suggesting that it may be more of a 'visual specialist' than the DFB or WBBB, for which visual responses may be more contingent on olfactory inputs.     

T-DNA标签在植物基因克隆和功能分析中的应用

在植物功能基因组学的研究中,插入突变已成为迅速识别和研究标签基因的一个有效遗传工具.本文介绍了T-DNA标签的概念及应用前提,详细论述了T-DNA标签在大规模植物基因功能分析中的应用以及使用启动子和增强子诱捕技术分离时空特异性启动子和表达基因,另外还分析了利用其特殊形式激活标签进行基因克隆和功能分析的优越性,并展望了T-DNA标签的应用前景.     

转基因白桦外源基因的多重PCR快速检测

根据转化的载体序列上T-DNA中的目的基因bt,选择性筛选标记基因nptⅡ和报告基因gus设计三对特异性引物,PCR产物片断大小分别为247、449、668 bp,应用多重PCR (mutiplex-PCR)的方法同步检测18株转基因白桦中三个基因的整合状况;用阳性对照为模板,对单重PCR（simplex-PCR）和多重PCR的各项指标进行比较。结果表明多重PCR检测多个外源基因在敏感性方面与单重PCR相比并没有减弱,而且略有提高;对18株样品的多重PCR同步检测无假阳性出现,结果准确,同时在操作中具有减少污染,缩短时间和节约成本等优点。因此,在对转基因白桦的外源基因的定期检测中,多重PCR是一种非常有效而便捷的方法,可以为转基因的拷贝数,T-DNA旁侧序列特征等转基因整合特性方面的研究提供数据。     

Number and Accuracy of T-DNA Insertions in Transgenic Banana (Musa spp.) Plants Characterized by an Improved Anchored PCR Technique

Nineteen transgenic banana plants, produced via Agrobacterium-mediated transformation, were analyzed for the integration of T-DNA border regions using an improved anchored PCR technique. The method described is a relatively fast, three-step procedure (restriction digestion of genomic DNA, ligation of ‘vectorette’-type adaptors, and a single round of suppression PCR) for the amplification of specific T-DNA border-containing genomic fragments. Most transgenic plants carried a low number of inserts and the method was suitable for a detailed characterization of the integration events, including T-DNA border integrity as well as the insertion of non-T-DNA vector sequences, which occurred in 26% of the plants. Furthermore, the particular band pattern generated by four enzyme/primer combinations for each individual plant served as a fingerprint, allowing the identification of plants representing identical transformation events. Genomic Southern hybridization and nucleotide sequence analysis of amplification products confirmed the data obtained by anchored PCR. Sequencing of seven right or left border junction regions revealed different T-DNA processing events for each plant, indicating a relatively low frequency of precisely nicked T-DNA integration among the plants studied.     

Generation of a flanking sequence-tag database for activation-tagging lines in japonica rice

We have generated 47,932 T-DNA tag lines in japonica rice using activation-tagging vectors that contain tetramerized 35S enhancer sequences. To facilitate use of those lines, we isolated the genomic sequences flanking the inserted T-DNA via inverse polymerase chain reaction. For most of the lines, we performed four sets of amplifications using two different restriction enzymes toward both directions. In analyzing 41,234 lines, we obtained 27,621 flanking sequence tags (FSTs), among which 12,505 were integrated into genic regions and 15,116 into intergenic regions. Mapping of the FSTs on chromosomes revealed that T-DNA integration frequency was generally proportional to chromosome size. However, T-DNA insertions were non-uniformly distributed on each chromosome: higher at the distal ends and lower in regions close to the centromeres. In addition, several regions showed extreme peaks and valleys of insertion frequency, suggesting hot and cold spots for T-DNA integration. The density of insertion events was somewhat correlated with expressed, rather than predicted, gene density along each chromosome. Analyses of expression patterns near the inserted enhancer showed that at least half the test lines displayed greater expression of the tagged genes. Whereas in most of the increased lines expression patterns after activation were similar to those in the wild type, thereby maintaining the endogenous patterns, the remaining lines showed changes in expression in the activation tagged lines. In this case, ectopic expression was most frequently observed in mature leaves. Currently, the database can be searched with the gene locus number or location on the chromosome at http://www.postech.ac.kr/life/pfg/risd. On request, seeds of the T(1) or T(2) plants will be provided to the scientific community.     

DIFFERENCES IN THE TIMING OF PRECHONDROGENIC LIMB DEVELOPMENT IN MAMMALS: THE MARSUPIAL–PLACENTAL DICHOTOMY RESOLVED

In contrast to placentals, marsupials are born with forelimbs that are greatly developmentally advanced relative to their hind limbs. Despite significant interest, we still do not know why this is the case, or how this difference is achieved developmentally. Studies of prechondrogenic and chondrogenic limbs have supported the traditional hypothesis that marsupial forelimb development is accelerated in response to the functional requirements of the newborn's crawl to the teat. However, limb ossification studies have concluded that, rather than the forelimb being accelerated, hind limb development is delayed. By increasing the taxonomic coverage and number of prechondrogenic events relative to previous studies, and combining traditional phylogenetic analyses of event sequences with novel analyses of relative developmental rates, this study demonstrates that the timing of limb development in marsupials is more complex than commonly thought. The marsupial phenotype was derived through two independent evolutionary changes in developmental rate: (1) an acceleration of the forelimb's first appearance and (2) a delay of hind limb development from the bud stage onward. Surprisingly, this study also provides some support for an evolutionary acceleration of the marsupial hind limb's first appearance. Further study is needed on the developmental and genetic mechanisms driving these major evolutionary transitions.     

Separation technologies for stem cell bioprocessing

Stem cells have been the focus of an intense research due to their potential in Regenerative Medicine, drug discovery, toxicology studies, as well as for fundamental studies on developmental biology and human disease mechanisms. To fully accomplish this potential, the successful application of separation processes for the isolation and purification of stem cells and stem cell‐derived cells is a crucial issue. Although separation methods have been used over the past decades for the isolation and enrichment of hematopoietic stem/progenitor cells for transplantation in hemato‐oncological settings, recent achievements in the stem cell field have created new challenges including the need for novel scalable separation processes with a higher resolution and more cost‐effective. Important examples are the need for high‐resolution methods for the separation of heterogeneous populations of multipotent adult stem cells to study their differential biological features and clinical utility, as well as for the depletion of tumorigenic cells after pluripotent stem cell differentiation. Focusing on these challenges, this review presents a critical assessment of separation processes that have been used in the stem cell field, as well as their current and potential applications. The techniques are grouped according to the fundamental principles that govern cell separation, which are defined by the main physical, biophysical, and affinity properties of cells. A special emphasis is given to novel and promising approaches such as affinity‐based methods that take advantage of the use of new ligands (e.g., aptamers, lectins), as well as to novel biophysical‐based methods requiring no cell labeling and integrated with microscale technologies. Biotechnol. Bioeng. 2012; 109: 2699–2709. © 2012 Wiley Periodicals, Inc.