Direct introduction of cloned DNA into the sea urchin zygote nucleus, and fate of injected DNA

A method is described for microinjection of cloned DNA into the zygote nucleus of Lytechinus variegatus. Eggs of this species are unusually transparent, facilitating visual monitoring of the injection process. The initial fate of injected DNA fragments appears similar to that observed earlier for exogenous DNA injected into unfertilized egg cytoplasm. Thus after end-to-end ligation, it is replicated after a lag of several hours to an extent indicating that it probably participates in most of the later rounds of DNA synthesis undergone by the host cell genomes during cleavage. The different consequences of nuclear versus cytoplasmic injection are evident at advanced larval stages. Larvae descendant from eggs in which exogenous DNA was injected into the nuclei are four times more likely (32% versus 8%) to retain this DNA in cell lineages that replicate very extensively during larval growth, i.e. the lineages contributing to the imaginal rudiment, and thus to display greatly enhanced contents of the exogenous DNA. Similarly, 36% of postmetamorphic juveniles from a nuclear injection sample retained the exogenous DNA sequences, compared to 12% of juveniles from a cytoplasmic injection sample. However, the number of copies of the exogenous DNA sequences retained per average genome in postmetamorphic juveniles was usually less than 0.1 (range 0.05-50), and genome blot hybridizations indicate that these sequences are organized as integrated, randomly oriented, end-to-end molecular concatenates. It follows that only a small fraction of the cells of the average juvenile usually retains the exogenous sequences. Thus, even when introduced by nuclear microinjection, the stable incorporation of exogenous DNA in the embryo occurs in a mosaic fashion, although in many recipients the DNA enters a wider range of cell lineages than is typical after cytoplasmic injection. Nuclear injection would probably be the route of choice for studies of exogenous DNA function in the postembryonic larval rudiment

An Ontology for Product-Service Systems

Industries are transforming their business strategy from a product-centric to a more service-centric nature by bundling products and services into integrated solutions to enhance the relationship between their customers. Since Product- Service Systems design research is currently at a rudimentary stage, the development of a robust ontology for this area would be helpful. The advantages of a standardized ontology are that it could help researchers and practitioners to communicate their views without ambiguity and thus encourage the conception and implementation of useful methods and tools. In this paper, an initial structure of a PSS ontology from the design perspective is proposed and evaluated

Negative spatial regulation of the lineage specific CyIIIa actin gene in the sea urchin embryo

The CyIIIa·CAT fusion gene was injected into Strongylocentrotus purpuratus eggs, together with excess ligated competitor sequences representing subregions of the CyIIIa regulatory domain. In this construct, the chloramphenicol acetyltransferase (CAT) reporter gene is placed under the control of the 2300 nucleotide upstream regulatory domain of the lineage-specific CyIIIa cytoskeletal actin gene. CAT mRNA was detected by in situ hybridization in serial sections of pluteus stage embryos derived from the injected eggs. When carrier DNA lacking competitor CyIIIa fragments was coinjected with CyIIIa.CAT, CAT mRNA was observed exclusively in aboral ectoderm cells, i.e. the territory in which the CyIIIa gene itself is normally expressed (as also reported by us previously). The same result was obtained when five of seven different competitor subfragments bearing sites of DNA-protein interaction were coinjected. However, coinjection of excess quantities of either of two widely separated, nonhomologous fragments of the CyIIIa regulatory domain produced a dramatic ectopic expression of CAT mRNA in the recipient embryos. CAT mRNA was observed in gut, mesenchyme cells and oral ectoderm in these embryos. We conclude that these fragments contain regulatory sites that negatively control spatial expression of the CyIIIa gene

A Grounded Theory of Sport Injury-Related Growth

Although previous research has shown that experiencing an injury can act as a catalyst for self-development, research that has examined the concept of sport injury-related growth (SIRG) remains largely descriptive. This study aimed to address this by developing a substantive theory to explain the processes through which injured athletes experienced SIRG. Using Strauss and Corbin’s (1998) variant of grounded theory, 37 injured athletes competing in a range of sports and competitive levels participated in qualitative interviews. Interviews (N=70) and data analysis were conducted over a period of 24 months. Transcripts were analyzed using open, axial, and selective coding. Quality criteria used were fit, relevance, workability, and modifiability. The grounded theory produced (i.e., Theory of Sport Injury-Related Growth) suggests a number of internal (i.e., personality, coping styles, knowledge and prior experience, and perceived social support) and external factors (i.e., cultural scripts, physical resources, time, and received social support) enable injured athletes to transform their injury into an opportunity for growth and development. The mechanisms through which this occurs are meta-cognitions, positive reappraisal, positive emotions, and facilitative responses. This theory offers a number of exciting avenues for future research, and provides medical personnel and practicing sport psychologists with a detailed explanation of how sport injury can lead to growth experiences

Competitive titration in living sea urchin embryos of regulatory factors required for expression of the CyIIIa actin gene

Previous studies have located some twenty distinct sites within the 2.3 kb 5' regulatory domain of the sea urchin CyIIIa cytoskeletal actin gene, where there occur in vitro high-specificity interactions with nuclear DNA-binding proteins of the embryo. This gene is activated in late cleavage, exclusively in cells of the aboral ectoderm cell lineages. In this study, we investigate the functional importance in vivo of these sites of DNA-protein interaction. Sea urchin eggs were coinjected with a fusion gene construct in which the bacterial chloramphenicol acetyltransferase (CAT) reporter gene is under the control of the entire CyIIIa regulatory domain, together with molar excesses of one of ten nonoverlapping competitor subfragments of this domain, each of which contains one or a few specific site(s) of interaction. The exogenous excess binding sites competitively titrate the available regulatory factors away from the respective sites associated with the CyIIIa.CAT reporter gene. This provides a method for detecting in vivo sites within the regulatory domain that are required for normal levels of expression, without disturbing the structure of the regulatory domain. We thus identify five nonoverlapping regions of the regulatory DNA that apparently function as binding sites for positively acting transcriptional regulatory factors. Competition with a subfragment bearing an octamer site results in embryonic lethality. We find that three other sites display no quantitative competitive interference with CyIIIa.CAT expression, though as shown in the accompanying paper, two of these sites are required for control of spatial expression. We conclude that the complex CyIIIa regulatory domain must assess the state of many distinct and individually necessary interactions in order to properly regulate CyIIIa transcriptional activity in development

Spin wave excitations in exchange biased IrMn/CoFe bilayers

Using an atomistic spin model, we have simulated spin wave injection and propagation into antiferromagnetic IrMn from an exchange coupled CoFe layer. The spectral characteristics of the exited spin waves have a complex beating behavior arising from the non-collinear nature of the antiferromagnetic order. We find that the frequency response of the system depends strongly on the strength and frequency of oscillating field excitations. We also find that the strength of excited spin waves strongly decays away from the interfacial layer with a frequency dependent attenuation. Our findings suggest that spin waves generated by coupled ferromagnets are too weak to reverse IrMn in their entirety even with resonant excitation of a coupled ferromagnet. However, efficient spin wave injection into the antiferromagnet is possible due to the non-collinear nature of the IrMn spin ordering

Micromagnetic modeling of the heat-assisted switching process in high anisotropy FePt granular thin films

The dynamic process of assisted magnetic switchings has been simulated to investigate the associated physics. The model uses a Voronoi construction to determine the physical structure of the nanogranular thin film recording media, the Landau-Lifshitz-Bloch equation is solved to evolve the magnetic system in time. The reduction of the magnetization is determined over a range of peak system temperatures and for a number of anisotropy values. The results show that the heat-assisted magnetic recording process is not simply magnetization reversal over a thermally reduced energy barrier. To achieve full magnetization reversal (for all anisotropies investigated), an applied field strength of at least 6 kOe is required and the peak system temperature must reach at least the Curie point (T c). When heated to T c, the magnetization associated with each grain is destroyed, which invokes the non-precessional linear reversal mode. Reversing the magnetization through this linear reversal mode is favorable, as the reversal time is two orders of magnitude smaller than that associated with precession. Under these conditions, as the temperature decreases to ambient, the magnetization recovers in the direction of the applied field, completing the reversal process. Also, the model produces results that are consistent with the concept of thermal writability; when heating the media to T c, the smaller grains require a larger field strength to reverse the magnetization