Analysis of mitochondrial control region DNA variation in New Zealand's brushtail possums (Trichosurus vulpecula) : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Ecology at Massey University

Brushtail possums (Trichosurus vulpecula) were first introduced from Australia to New Zealand in 1858 to establish a fur industry. Currently numbering more than 65 million, they are recognised as the most important mammalian pest in New Zealand, because of the environmental and agricultural damage they cause. Possums act as a wildlife reservoir of bovine tuberculosis (Tb) and, as such, threaten New Zealand's multi-million dollar beef and dairy industry. Eliminating bovine Tb in livestock requires removal of contact with infected possums. This is mainly achieved through the intensive poisoning of areas of known wildlife Tb infection and the establishment around them of zones of low possum density (known as buffer zones) adjacent to at-risk farmland. Not only does this result in lower possum density, and thus fewer dispersing possums, but may also affect the movement patterns of possums. Measurement of gene frequency differences between populations associated with a buffer zone would allow a qualitative estimate of the effect of buffer zones on limiting possum movement. The mitochondrial DNA (mtDNA) control region is an effective marker for detecting intraspccific genetic structure because it has a high mutation rate, lack of recombination and uniparental mode of inheritance. An extensive survey of brushtail possum mtDNA control region variation in New Zealand was conducted to quantify levels of variation and thus assess the utility of the mtDNA control region as a marker for detecting genetic differentiation between possum populations. Nine haplotypes were found among 70 possums from throughout New Zealand. Most of the variation (six haplotypes) was concentrated in the North Island, and the most widespread haplotype (occurring in all four islands surveyed) was also the most common - found in 67% of possums surveyed. The technique of single stranded conformation polymorphism (SSCP) was developed for the brushtail possum so that a quick, cost-effective and sensitive method for surveying mtDNA control region variation in large numbers of individuals was available. This assay was applied to screen the variation in possums separated by small spatial scales associated with two buffer zones in the South Island. A total of 234 possums were screened, with 98.7% found to possess the same haplotype. The other 1.3%, all from one location, possessed a second haplotype. The extremely low levels of variation makes it highly unlikely that surveys of variation in mtDNA will be able to detect an effect of buffer zones on possum movement, at least in the South Island. Areas of higher variation, such as certain parts on the North Island, would be better candidates for testing the effect of barriers such as buffer zones on genetic differentiation between possum populations

P53 tumour-suppressor gene mutations are mainly localised on exon 7 in human primary and metastatic prostate cancer.

Mutations in the p53 tumour-suppressor gene are among the most common genetic alterations in human cancers. In the present study we analysed the mutations in the p53 tumor-suppressor gene in 25 primary and 20 metastatic human prostate cancer specimens. DNA extracted from the paraffin-embedded sections was amplified by hot-start polymerase chain reaction, and p53 gene mutations in the conserved mid-region (exons 4-9) were examined using single-strand conformation polymorphism (SSCP) analysis and immunohistochemistry. In the present study, we used a novel hot-start PCR-SSCP technique using DNA Taq polymerase antibody, which eliminates primer-dimers and non-specific products. Because of this new technique, the results of PCR-SSCP showed very high resolution. Polymerase chain reaction products were sequenced directly for point mutations for the p53 gene. Mutations were found in 2 out of 25 primary prostate cancers (8%) and 4 out of 20 metastatic cancers (20%). Mutations were observed exclusively in exon 7 and not in exons 4, 5, 6, 8 or 9. Nuclear accumulation of p53 protein, determined by immunohistochemistry, correlated with the degree of metastasis in prostatic cancer

Discrete Scale Relativity

The possibility that global discrete dilation invariance is a fundamental symmetry principle of nature is explored. If the discrete self-similarity observed in nature is exact, then the Principle of General Covariance needs to be broadened in order to accommodate this form of discrete conformal invariance, and a further generalization of relativity theory is required.Comment: 9 pages, minor revisions, accepted at Astrophys. Space Sci., comments welcom

Starch and oil in the donor cow diet and starch in substrate differently affect the in vitro ruminal biohydrogenation of linoleic and linolenic acids

Trans isomers of fatty acids exhibit different health properties. Among them, trans-10,cis-12 conjugated linoleic acid has negative effects on milk fat production and can affect human health. A shift from the trans-11 to the trans-10 pathway of biohydrogenation (BH) can occur in the rumen of dairy cows receiving high-concentrate diets, especially when the diet is supplemented with highly unsaturated fat sources. The differences of BH patterns between linoleic acid (LeA) and linolenic acid (LnA) in such ruminal conditions remain unknown; thus, the aim of this work was to investigate in vitro the effects of starch and sunflower oil in the diet of the donor cows and starch level in the incubates on the BH patterns and efficiencies of LeA and LnA. The design was a 4 × 4 Latin square design with 4 cows, 4 periods, and 4 diets with combinations of 21 or 34% starch and 0 or 5% sunflower oil. The rumen content of each cow during each period was incubated with 4 substrates, combining 2 starch levels and either LeA or LnA addition. Capillary electrophoresis single-strand conformation polymorphism of incubates showed that dietary starch decreased the diversity of the bacterial community and the high-starch plus oil diet modified its structure. High-starch diets poorly affected isomerization and first reduction of LeA and LnA, but decreased the efficiencies of trans-11,cis-15-C18:2 and trans C18:1 reduction. Dietary sunflower oil increased the efficiency of LeA isomerization but decreased the efficiency of trans C18:1 reduction. An interaction between dietary starch and dietary oil resulted in the highest trans-10 isomers production in incubates when the donor cow received the high-starch plus oil diet. The partition between trans-10 and trans-11 isomers was also affected by an interaction between starch level and the fatty acid added to the incubates, showing that the trans-10 shift only occurred with LeA, whereas LnA was mainly hydrogenated via the more usual trans-11 pathway, whatever the starch level in the substrate, although the bacterial communities were not different between LeA and LnA incubates. In LeA incubates, trans-10 isomer production was significantly related to the structure of the bacterial community

A novel octopamine receptor with preferential expression in <i>Drosophila</i> mushroom bodies

Octopamine is a neuromodulator that mediates diverse physiological processes in invertebrates. In some insects, such as honeybees and fruit flies, octopamine has been shown to be a major stimulator of adenylyl cyclase and to function in associative learning. To identify an octopamine receptor mediating this function in Drosophila, putative biogenic amine receptors were cloned by a novel procedure using PCR and single-strand conformation polymorphism. One new receptor, octopamine receptor in mushroom bodies (OAMB), was identified as an octopamine receptor because human and Drosophila cell lines expressing OAMB showed increased cAMP and intracellular Ca2+ levels after octopamine application. Immunohistochemical analysis using an antibody made to the receptor revealed highly enriched expression in the mushroom body neuropil and the ellipsoid body of central complex, brain areas known to be crucial for olfactory learning and motor control, respectively. The preferential expression of OAMB in mushroom bodies and its capacity to produce cAMP accumulation suggest an important role in synaptic modulation underlying behavioral plasticity