The development of property rights in biotechnology

The translation of biological theory into engineering (biotechnology) has resulted in the development of novel products and processes. Some of these products are living organisms, usually containing unique genetic arrangements not found in nature. The extension of legal protection to products and processes was required in order for biotechnology to become an unexceptional way in which to do business. The American experience with biotechnology, repeated elsewhere, is demonstrated to have proceeded first through the negotiation of obstacles in administrative law and second through challenges to property law. This outcome for the regulatory management of biotechnology and the legal protection of its products may be interpreted as a function of a cultural bias for scientific authority and progress ideology.     

The macrolepidoptera of farm woodlands: determinants of diversity and community structure

The Farm Woodland Scheme, which provided incentives to convert agricultural land to timber production, contained an implicit assumption that farm woodlands produce important benefits for wildlife. The moth fauna of 18 farm woodlands in the Vale of York was surveyed between May and November 1991. The aims were twofold. The first was to determine if there were benefits for moth species diversity. The second was to ascertain whether concepts of island biogeography and the plant species richness of the woods were related to the moth species composition.Eleven families, 214 species and over 16 000 individuals of moths were recorded. Classification of the species presence/absence matrix indicated that small woods (less than 1ha) did not have characteristic woodland moth communities. Woods larger than 5ha were judged to be more valuable for the long-term conservation of woodland moth diversity. The best predictor of moth species richness was the herbaceous plant species richness within woodlands. Species richness of the family Geometridae was positively related to woodland area, as well as to woodland shape (compact shapes being preferable to elongated shapes). Characteristic woodland species are influenced by isolation (less isolated woods being richer in species). The implications of different powers of dispersal between moth families are discussed. Farm woodlands will be of more value for the conservation of the Macrolepidoptera if they are large, compact and incorporate remnants of existing woodland with extant herbaceous vegetation. These should be factors which are taken into consideration when providing incentives to establish and manage farm woodlands.     

Biodiversity in agricultural landscapes: the ground beetle communities of woody uncultivated habitats

Agricultural landscapes can be defined as mosaics of landscape elements which are affected by farming practices. Woodland habitats, even though they are managed, are amongst the most stable elements of agricultural landscapes and can play a key role in the maintenance of biodiversity. This study of the ground beetle (carabid) communities of woodlands and woody linear features in a Scottish agricultural landscape shows that these habitats contribute significantly to the overall landscape diversity of these beetles. Communities in woods and hedgerows display the same species diversity and are both characterized by the presence of forest species. The main factors constraining carabid communities in both environments are the grazing intensity and, to a lesser extent, the type of soil. Heavily grazed locations are characterized by the occurrence of grassland species while forest species are restricted to ungrazed locations. At the landscape scale, the distribution of the forest species is limited by spatial isolation, indicating that there are insufficient functional links between woodland habitats in the study area. Isolation could be compensated for either by a better control of grazing so that linear features can be used as dispersal corridors for forest carabids or by planting more linear features and woods in the area.     

Mitochondrial and Peroxisomal β-Oxidation of Stearic and Lignoceric Acids by Rat Brain

Crude subcellular fractions were prepared from adult rat brains by differential centrifugation of brain homogenates. Greater than 98% of the cellular mitochondrial marker enzyme activity sedimented in the heavy and light mitochondrial pellets, and less than 1% of the activity sedimented in microsomal pellets. Lysosomal marker enzyme activities mainly (71-78% of cellular activity) sedimented in the heavy and light mitochondrial pellets. Significant amounts of the lysosomal marker enzyme activity also sedimented in the crude microsomal pellets (9-13% of total) and high-speed supernatants (14-16% of total). The specific activities of microsomal and peroxisomal marker enzyme activities were highest in the crude microsomal pellets. Fractionation of the crude microsomal pellets on Nycodenz gradients resulted in the separation of the bulk of the remaining mitochondrial, lysosomal, and microsomal enzyme activities from peroxisomes. Fatty acyl-CoA synthetase activities separated on Nycodenz gradients as two distinct peaks, and the minor peak of the activities was in the peroxisomal enriched fraction. Fatty acid beta-oxidation activities also separated as two distinct peaks, and the activities were highest in the peroxisomal enriched fractions. Mitochondria were purified from the heavy mitochondrial pellets by Percoll density gradients. Fatty acyl-CoA synthetase and fatty acid beta-oxidation activities were present in both the purified mitochondrial and peroxisomal enriched fractions. Stearoyl-CoA synthetase activities were severalfold greater compared to lignoceroyl-CoA synthetase, and stearic acid beta-oxidation was severalfold greater compared to lignoceric acid beta-oxidation in purified mitochondrial and peroxisomal enriched fractions.(ABSTRACT TRUNCATED AT 250 WORDS)     

Oral dosing of insects with feeding deterrent compounds

A preliminary assessment was made of a variety of approaches to dosing insects with compounds that normally deter feeding. Enclosing water-soluble compounds in liposomes has promise for strong deterrents, although in drinking assays many deterrents lose their potency anyway. Microencapsulating compounds by embedding them in a polymer matrix to form spheres without a true barrier wall may have masked deterrency of a lipophilic compound, but did not adequately mask the taste of water-soluble compounds. Microencapsulation with gelatin was shown to work with lipid-soluble dye and the capsule walls appear to be broken down very rapidly after ingestion. Coating fine particles with beeswax did not successfully mask the deterrence of quinine, but has value with compounds insoluble in lipids. Starch coating can be effective for insects lacking salivary amylase. Molecular encapsulation of grindelic acid in cyclodextrin masked its deterrency to Schistocerca americana. Nymphs fed this material through the second and third instars showed an increase in development time during the third instar without any corresponding reduction in consumption rate. Behavioral approaches to administering deterrent compounds include drinking and habituation. Some compounds that reduce consumption of leaf material were taken in drinking water in amounts sufficient to give relevant doses in one or two drinks daily. Pseudaletia unipuncta larvae fed nicotine-treated wheat through the fifth and sixth instars ate less than control larvae during the first 24 h of the experiment but showed no reduction in consumption after that period. A one-day increase in development time was attributed to the one-day habituation period. More work is needed on the release of taste-masked compounds in the gut.

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Resumé Des essais préliminaires ont porté sur les différentes techniques de dosage de produits qui normalement inhibent la prise de nourriture. L'isolement des composés hydrosolubles dans des liposomes a donné des résultats prometteurs pour les dissuadants puissants, bien qu'au cours d'absorptions expérimentales de liquide beaucoup de dissuadants aient perdu de leur efficacité. La microencapsulation, en les enfouissant dans une matrice de polymère pour former des sphères sans véritable paroi, peut avoir masqué le pouvoir dissuadant des produits lipophiles, mais n'a pas masqué suffisamment le goût des produits hydrosolubles. La microencapsulation dans des capsules de gélatine a été efficace avec les colorants liposolubles, la paroi de la capsule semblant détruite très rapidement après ingestion. Le revêtement de fines particules par de la cire n'a pas masqué efficacement l'effet dissuasif de la quinine, mais a été valable avec les produits insolubles dans les lipides. L'encapsulation moléculaire de l'acide grindellique dans de la cyclodextrine a masqué son effect dissuasif pour [?]Schistocerca gregaria. Les larves ont mangé le tout pendant les second et troisième stades et ont présenté un ralentissement du développement pendant le troisième stade sans diminuation de la consommation. L'étude comportementale de l'administration de produits dissuasifs comprend l'action de boire et l'accoutumance. Certains produits qui réduisent la consommation de feuilles ont été absorbés en quantités suffisantes avec l'eau de boisson pour donner des doses significatives avec une ou deux prises de liquide par jour. Des larves de Pseudaletia unipuncta alimentées avec du blé traité à la nicotine pendant les cinquième et sixième stades, ont consommé moins que les témoins, mais seulement pendant les 24 premières heures de l'expérience. La prolongation d'un jour, de la durée de développement, a été attribuée au temps nécessaire à l'accoutumance.

     

Behaviour of tritium-labelled isopenicillin N and 6-aminopenicillanic acid as potential penicillin precursors in an extract of Penicillum chrysogenum.

1. 3H was incorporated into solvent-soluble penicillin from isopenicillin N and 6-aminopenicillanic acid 3H-labelled in the 2beta-methyl group when the labelled compounds were incubated with a crude extract of Penicillum chrysogenum. 2. With a soluble protein fraction of the extract incorporation from isopenicillin N occurred on addition of phenyl-acetyl-CoA. 3. Labelled benzylpenicillin was isolated after incubation of the crude extract with phenylacetyl-CoA and isopenicillin and the addition of unlabelled benzylpenicillin as a carrier. 4. No incorporation of 3H into solvent-soluble penicillin was detected on incubation of these extracts with penicillin N.     

Geometry of the dry-state oligomerization of 2′,3′-cyclic phosphates

Summary Evaporation of a solution of thymidine plus either theexo or theendo diastereomer of uridine cyclic 2,3-O, O-phosphorothioate (U > p(S) in 1,2-diaminoethane hydrochloride buffer gave the 2,5 and 3,5 isomers of (P-thio) uridylylthymidine (Up(S)dT) in a ratio of 1:2 with a combined yield of about 20%. These isomers were re-converted to U > p(S) and dT by a reaction that is known to proceed by an in-line mechanism. Both the 2,5 and 3,5 isomers gave as product the same diastereomer of U > p(S) that had been used originally in their formation. These dry-state prebiotic reactions (Verlander, Lohrmann, and Orgel 1973) are thus shown to be stereospecific, and both the 2,5 and 3,5 internucleotide bonds are formed by an in-line mechanism.Abbreviations DAE 1,2-diaminoethane - HPLC high pressure liquid chromatography - RNase bovine pancreatic ribonuclease A, EC 3.1.4.22 - TEAB triethylammonium bicarbonate - tris tris(hydroxymethyl)aminomethane - UMP(S) uridine monophosphorothioate - U > p uridine cyclic 2,3-phosphate - U > p(S) uridine cyclic 2,3-O, O-phosphorothioate - Up(S)dT (P-thio)uridylylthymidine - U2p(R_p-S)5dT (P-thio)uridylylthymidine with theR configuration at phosphorous, and a 2,5 internucleotide linkage     

Muscle development in Atlantic salmon (Salmo salar) embryos and the effect of temperature on muscle cellularity

Salmon eggs were incubated at 5, 8 or 11° C from fertilization to hatching. At Gorodilov stages 25, 27, 29, 31 and 33 transverse sections of whole embryos (at somite level 10–15) were prepared for histochemistry and electron microscopy. At every stage up to hatching, cross–sectional areas of the embryos were not different between temperatures, and from stage 27 onwards there was also no difference in the ratio of white to red muscle. However, there were more muscle fibres but of smaller average diameter in both the red and white muscle for the colder temperature embryos. At hatching there were also more nuclei (per cross–section) in the colder embryos but more nuclei per muscle fibre in the warmer embryos. In all cases the 8° C embryos were intermediate between 5 and 11° C embryos in their muscle parameters. Fast and slow muscle fibres could only be distinguished in the embryos by alkali–stable ATPase reactions. Succinic dehydrogenase activity was low in embryonic fish. No differences between the temperature groups were detected in the histochemical reactions for either ATPase or succinic dehydrogenase activities.     

Recombination between Homoeologous Chromosomes of Lager Yeasts Leads to Loss of Function of the Hybrid GPH1 Gene

Yeasts used in the production of lagers contain complex allopolyploid genomes, resulting from the fusion of two different yeast species closely related to Saccharomyces cerevisiae and Saccharomyces bayanus. Recombination between the homoeologous chromosomes has generated a number of hybrid chromosomes. These recombination events provide potential for adaptive evolution through the loss or gain of gene function. We have examined the genotypic and phenotypic effects of one of the conserved recombination events that occurred on chromosome XVI in the region of YPR159W and YPR160W. Our analysis shows that the recombination event occurred within the YPR160W gene, which encodes the enzyme glycogen phosphorylase and generates a hybrid gene that does not produce mature mRNA and is nonfunctional due to frameshifts in the coding region. The loss of function of the hybrid gene leads to glycogen levels similar to those found in haploid yeast strains. The implications for the control of glycogen levels in fermentative yeasts are discussed.Yeasts used in the production of lagers, originally referred to as Saccharomyces carlsbergensis, are now classified as Saccharomyces pastorianus (18). Lager yeasts contain complex polyploid genomes with general tetraploid DNA content and are believed to have arisen from a natural fusion of two different haploid species followed by a genome duplication event or alternatively from the fusion of two diploid yeast species (2, 4, 14, 18). Subsequent genome changes, such as chromosome loss and/or duplication and translocations, have resulted in unequal numbers of chromosomes in the present-day strains, a state referred to as aneuploidy.A sequence analysis of individual genes indicated that the parental strains contributing to the hybrid strain closely resemble Saccharomyces cerevisiae and Saccharomyces bayanus (6, 15, 16). A whole-genome sequence analysis of the lager yeast strain Weihenstephan (15) identified the presence of three types of chromosomes, referred to as (i) S. cerevisiae-like chromosomes, (ii) S. bayanus-like chromosomes, and (iii) hybrids resulting from recombination events between the homoeologous parental chromosomes.Competitive genomic hybridization (CGH) analysis of two S. pastorianus strains, named CMBS-33 and 6701, identified as many as 28 specific locations where recombination between homoeologous pairs of chromosomes or chromosomal translocations may have occurred (3). Of the 28 sites identified, 13 occur at unique sites on eight different chromosomes, while the rest are in subtelomeric X elements or within 25 kbp of the telomere. Many of the genes adjacent to the recombination sites encode proteins that play essential roles in fermentation, including ADH2, ADH4, AAD6 and TDH2 (ethanol metabolism), FLO10, and PHD1 (3).We have recently shown that recombination at these “hot spots” can be induced by the exposure of lager yeasts to environmental stresses such as high temperature and high osmotic stress (13). Furthermore, fermentation under stress conditions leads to the amplification and loss of telomeric regions on a selected set of chromosomes and gene amplification radiating from the rRNA locus on chromosome XII and the DUP locus on chromosome I (13). Since a number of genes, including the MAL (maltose utilization) and the FLO (flocculation) genes, encoding proteins required for the fermentation process reside at the telomeres, such genome dynamics can have important consequences for the immediate quality and outcome of fermentation in addition to severe consequences on strain stability and purity.One of the recombination events identified by CGH analysis is located on chromosome XVI in the region of YPR159W and YPR160W. DNA to the left of the region hybridizes to S. cerevisiae microarrays, while genes between YPR160W and YPR190C and encompassing approximately 58 kb of DNA displayed a lack of hybridization to these microarrays, suggestive of a hybrid chromosome (3). Whole-genome sequence analysis of the Weihenstephan strain confirmed the existence of hybrid chromosome XVI and indicated the presence a second type of chromosome XVI containing S. bayanus-like sequences to the left of YPR159W (15, 16).To examine the genotypic and phenotypic outcomes of this recombination event, the right arm of chromosome XVI has been cloned from the yeast strain CMBS-33. Our analysis reveals that the recombination event occurred within the open reading frame (ORF) of YPR160W (GPH1) encoding the enzyme glycogen phosphorylase, which is required for the mobilization of stored glycogen through its conversion into glucose-1-P. The recombination event generates a hybrid gene that does not produce a mature mRNA and is nonfunctional due to frameshifts in the coding region.     

Characterization of anti-TIMP-1 monoclonal antibodies for immunohistochemical localization in formalin-fixed, paraffin-embedded tissue.

The aim of this study was to evaluate seven anti-TIMP-1 (tissue inhibitor of metalloproteinase-1) monoclonal antibodies by immunohistochemical (IHC) staining of formalin-fixed, paraffin-embedded (FFPE) tissue. Detection of the TIMP-1 protein was studied by IHC in FFPE human archival normal and neoplastic samples. Indirect IHC technique was used, and the seven antibodies (clones VT1, VT2, VT4, VT5, VT6, VT7, and VT8) were tested in various concentrations using different pretreatment protocols. All seven VT antibodies specifically immunostained the cytoplasm of islets of Langerhans cells in normal pancreas, epithelial cells of hyperplastic prostate, tumor cells of medullary thyroid carcinoma, and fibroblast-like cells of malignant melanoma. Specificity of the anti-TIMP-1 antibodies was confirmed by several controls, e.g., Western blotting on proteins extracted from FFPE tissue showed that the VT7 antibody reacted specifically with a protein band of approximately 28 kDa, corresponding to the molecular mass of TIMP-1. However, sensitivity varied with the different antibodies. Use of heat-induced epitope retrieval (HIER) and the VT7 clone applied at low concentrations demonstrated more intense immunoreactivity with the TIMP-1-positive cell types compared to the other six clones. Furthermore, when tested on a range of normal and neoplastic endocrine tissues, the VT7 clone demonstrated immunoreactivity with all neuroendocrine cell types. In conclusion, all seven antibodies detected TIMP-1 protein in various normal and neoplastic FFPE tissues, but one clone, VT7, was superior for IHC staining of TIMP-1 in FFPE tissue sections when using HIER.