Investigation of Ochratoxin A biosynthetic genes in<Emphasis Type="Italic">Penicillium verrucosum</Emphasis> by DDRT-PCR experiments: differential expression of OTA genes

A defined minimal medium has been developed which is either restrictive or permissive for the production of ochratoxin A byPenicillium verrucosum simply by changing the nitrogen and carbon source. The combination of ammonium/glycerin promoted, whereas the combination nitrate/glucose repressed ochratoxin production.In parallel mutants ofP verrucosum have been constructed by restriction endonuclease mediated integration, which were not able to produce ochratoxin. Different types of transformants, which either produce no detectable amounts of ochratoxin A but ochratoxin α, strains which produce only ochratoxin B and strains which produces none of these secondary metabolites, have been observed.     

Structural comparison of fibroblast growth factor-specific heparan sulfates derived from a growing or differentiating neuroepithelial cell line

Heparan sulfate (HS) glycosaminoglycans are essential modulators of fibroblast growth factor (FGF) activity both in vivo and in vitro, and appear to act by cross-linking particular forms of FGF to appropriate FGF receptors. We have recently isolated and characterized two separate HS pools derived from immortalized embryonic day 10 mouse neuroepithelial 2.3D cells: one from cells in log growth phase, which greatly potentiates the activity of FGF-2, and the other from cells undergoing contact-inhibition and differentiation, which preferentially activates FGF-1. These two pools of HS have very similar functional activities to those species isolated from primary neuroepithelial cells at corresponding stages of active proliferation or differentiation. We present here a structural comparison between these cell line HS species to establish the nature of the changes that occur in the biosynthesis of HS. A combination of chemical and enzymatic cleavage, low pressure chromatography and strong anion-exchange HPLC were used to generate full chain models of each species. Overall, the HS pools synthesized in the dividing cell line pools possessed less complex sulfation than those derived from more differentiated, growth arrested cells.      

Heterologous expression of an engineered truncated form of human Lewis fucosyltransferase (Fuc-TIII) by the methylotrophic yeast Pichia pastoris

A stable GS115 Pichia pastoris recombinant strain was constructed to secrete a truncated form of the human alpha(1,3/4) fucosyltransferase (amino acids 45-361). Enzyme production resulted from a secretory pathway based on the pre-pro- alpha mating factor signal sequence of the yeast Saccharomyces cerevisiae . Following its transit through the Golgi apparatus, the enzyme accumulated in the periplasmic space before its release in the culture broth (about 30 mg/l). Cell-enclosed enzyme ( approximately 0.16%) proved to be fairly stable for many freezing and thawing cycles and could be used several times as an immobilized catalyst. Soluble enzyme (>99.8%) representing the main protein of the culture broth (10%) has been characterized by Western-blotting, substrate specificities and kinetic parameters. The two forms (cell- enclosed and soluble) of recombinant enzyme may be used for in vitro synthesis of Lewisadeterminants.      

Development of Sporangiophores of Peronospora parasitica (Pers. ex Fr.) Fr.

The morphology of developing sporangiophores of Peronosporaparasitica from wallflower is described, and morphogenesis maybe divided into the following five stages: the sporangiophoreprimordium, unbranched sporangiophore, branched sporangiophore,spore formation and maturation, and formation of the cross wall.The growth of individual sporangiophores in a humidity chamberwas followed under the microscope, and increase in height andincrease in volume measured. The greatest increase in volumewas during spore formation, when the sporangiophore volume mightquadruple within an hour.     

Patterns of molecular genetic variation in Plantago major and P. intermedia in relation to ozone resistance

Patterns of molecular genetic variation were examined in seed collections of Plantago major and Plantago intermedia , used previously to investigate the variations in ozone (O₃) resistance of these species across Europe. Total genomic DNA was amplified with random primers (random amplied polymorphic DNA (RAPD) and inter- single sequence repeats (SSR)) to produce 73 genetic markers. In addition, allozyme and chloroplast variations were surveyed. Genetic markers were examined for association with O₃ resistance in 18 British populations of P. major as well as 27 continental European populations of P. major and P. intermedia . Two populations that exhibited increased resistance to O₃ following several years' exposure to high O₃ concentrations in the field showed decreased genetic variation over time. In addition, their genetic composition showed no drastic change, which suggests that the change in resistance to O₃ was probably the result of selection on genotypes already present in local populations (selection in situ ). It appears that selection for O₃ resistance may occur in independent populations, and also may involve a number of genetically determined traits. Consequently the finding that plants with similar degrees of O₃ resistance are not closely related was not unexpected. However, the finding of an association of several genetic markers with O₃ resistance merits further investigation.