Binding of phosphorothioate oligodeoxynucleotides to basic fibroblast growth factor, recombinant soluble CD4, laminin and fibronectin is P-chirality independent.

Antisense oligodeoxynucleotides can selectively inhibit the expression of individual genes and thus have potential applications in anticancer and antiviral therapy. A critical prerequisite to their use as therapeutic agents is the understanding of their non-specific interactions with biological structures, e.g. proteins. In this study we examined the interactions of P-chiral phosphorothioate oligodeoxynucleotides with several proteins. The Rp- and Sp- diastereomers, and racemic machine-made mixtures, or M-oligodeoxynucleotides were used independently as competitors of the binding of a probe, phosphodiester oligodeoxynucleotide bearing a 5' alkylating moiety, to rsCD4, bFGF and laminin. These oligodeoxynucleotides were also used as competitors of the binding of a non-alkylating probe M-phosphorothioate oligodeoxynucleotide, 5'-32P-SdT18 to fibronectin. The average values of and quantitative estimates for the IC50 of competition and the constant of competition (Kc) of Rp-, Sp- and M-stereoisomers of several homo- and heteropolymer oligodeoxynucleotides were determined and compared. Surprisingly, in the proteins we studied, the values of IC50 and Kc for the Rp-, Sp- and M-oligodeoxynucleotides were essentially identical. Thus, the ability of the phosphorothioate oligodeoxynucleotides we employed, to bind to the proteins studied in this work, is virtually independent of P-chirality. Our results also imply that the role of the purine and pyrimidine bases in oligodeoxynucleotide-protein interactions, as well as the nature of the contact points (sulfur versus oxygen) between the oligomer and the protein, may be relatively unimportant.     

The effects of temperature and the Rht3 dwarfing gene on growth, cell extension, and gibberellin content and responsiveness in the wheat leaf

The effects of low temperature and the Rht3 dwarfing gene onthe dynamics of cell extension in leaf 2 of wheat were examinedin relation to gibberellin (GA) content and GA-responsivenessof the extension zone. Leaf 2 of wild-type (rht3) wheat closelyresembled that of the Rht3 dwarf mutant when seedlings weregrown at 10C. The maximum relative elemental growth rate (REGR)within the extension zone in both genotypes was lower at 10Cthan at 20C, but the position with respect to the leaf basewas unaffected by temperature. The size of the extension zoneand epidermal cell lengths were similar in both genotypes at10C. Growth at 20C, instead of 10C, increased the lengthof the extension zone beyond the point of maximum REGR in thewild type, but not in the Rht3 mutant. Increasing temperatureresulted in longer epidermal cells in the wild type. Treatingwild-type plants at 10C with gibberellic acid (GA₃) also increasedthe length of the extension zone, but the Rht3 mutant was GA-non-responsive.However, the concentrations of endogenous GA₁ and GA₃ remainedsimilar across the extension zone of wild-type plants grownat both temperatures, despite large differences in leaf growthrates. The period of accelerating REGR as cells enter the extensionzone, and the maximum REGR attained, are apparently not affectedby GA. It is proposed that GA functions as a stimulus for continuedcell extension by preventing cell maturation in the region beyondmaximum REGR and that low temperature increases the sensitivitythreshold for GA action. Key words: Cell extension, gibberellin, Rht3 dwarfing gene, temperature, wheat leaf     

Acclimation to freezing temperatures in perennial ryegrass (Lolium perenne)

The increasing demands being placed on natural grasslands in the era following the appearance of Bovine Spongiform Encephalitis require that forage crops provide a reliable extended season of growth, combined with good winter survival to ensure sward longevity. The ability to tolerate sub-zero temperatures is integral to the survival of perennial forages. Since the development of freezing tolerance is crucial to the survival and productivity of over-wintering crops, forage breeding programmes require an improved understanding of the individual characteristics that contribute to tolerance to sub-zero temperatures. Photosynthesis, carbohydrate content and changes in protein composition were investigated in two varieties of Lolium perenne which differ in their response to growth at low temperature.     

Expression microarray hybridization kinetics depend on length of the immobilized DNA but are independent of immobilization substrate

Expression microarrays are often constructed by the immobilization of PCR products on two-dimensional modified glass slides or on three-dimensional microporous substrates. In this study we investigate whether the length of the immobilized species and the substrate choice influence hybridization dynamics. Using a simple bimolecular mass action controlled model to describe hybridization, we observed that the extent of hybridization and the initial velocities were directly dependent on the length of the immobilized species. An inflection point was noted at a length of 712 bases, above which the influence of length on hybridization rate decreased. Interestingly, we observed no differences in these parameters whether hybridization occurred on a two- or three-dimensional surface. Furthermore, the affinity of the solution phase labeled species for the immobilized species was identical for all arrayed lengths on both surfaces. These data indicate a similar interaction of the noncovalently immobilized species with either surface. Finally, we have determined that competitive hybridization on expression microarrays is nonlinear with respect to time and concentration of competitor. This observation is critical for analysis of expression array data.     

FAST slides: a novel surface for microarrays

We have evaluated FAST slides, a glass slide with a microporous polymeric surface that is a suitable substrate for microarray technology. The surface is a nitrocellulose-based polymer that binds DNA and proteins in a noncovalent but irreversible manner. FAST slides are compatible with robotic systems currently used to create microarrays and can easily accommodate volumes of 0.03-2 nL/spot. Our data indicate that FAST slides have a much higher binding capacity for DNA and better spot-to-spot consistency than traditional poly-lysine-coated slides. In addition, FAST slides are well suited for fluorescent detection because of their relatively low light scatter and efficient retention of arrayed DNA. These properties translate into fluorescent sensitivity comparable to modified glass surfaces. FAST slides are also ideal for arraying proteins, making them the only substrate of their kind currently available for microarray applications.     

Aboriginal ‘Difference’ and ‘Autonomy’ Then and Now: Four Decades of Change in a Western Desert Society

In just a few decades, Aboriginal people living near Australia's Western Desert fringe have experienced an extraordinarily intense trajectory of change: from a highly autonomous nomadic existence, through ‘first contacts’, the pastoral and mission frontiers, the devastating impacts of alcohol and of Western lifestyle diseases, the outstation movement, resource exploration and mining, a long but largely successful struggle for native title, and much else. In this paper, notions of ‘difference’ and ‘autonomy’ are used to explore these transformations. The situation among the Mardu is here linked to the gulf between government policies and lived Aboriginal experience. If the self‐management thrust of 1970s policies achieved partial restoration of Aboriginal autonomy, recent Federal Government policies are intent on intervention to reduce difference and claw back some of that autonomy. Their determination to force Aboriginal people out of their ‘dysfunctional’ ‘cultural museums’ (homeland settlements) and into greater economic engagement ignores the crucial underpinnings of security and identity among remote Aborigines. The retention of difference, albeit at considerable social cost and entrenched disadvantage, is still strongly preferred by Mardu to the kinds of engagement with the dominant society that not only assault their sense of self but also threaten to overwhelm whatever autonomy remains to them.