Methylation detection oligonucleotide microarray analysis: a high-resolution method for detection of CpG island methylation

Methylation of CpG islands associated with genes can affect the expression of the proximal gene, and methylation of non-associated CpG islands correlates to genomic instability. This epigenetic modification has been shown to be important in many pathologies, from development and disease to cancer. We report the development of a novel high-resolution microarray that detects the methylation status of over 25 000 CpG islands in the human genome. Experiments were performed to demonstrate low system noise in the methodology and that the array probes have a high signal to noise ratio. Methylation measurements between different cell lines were validated demonstrating the accuracy of measurement. We then identified alterations in CpG islands, both those associated with gene promoters, as well as non-promoter-associated islands in a set of breast and ovarian tumors. We demonstrate that this methodology accurately identifies methylation profiles in cancer and in principle it can differentiate any CpG methylation alterations and can be adapted to analyze other species.     

Genome‐wide analyses of rice root development QTLs and development of an online resource, Rootbrowse

Genetic control of root development in rice is complex and the underlying mechanisms (constitutive and adaptive) are poorly understood. Lowland and upland varieties of indica and japonica rice with contrasting root development characteristics have been crossed, mapping populations developed and a number of QTLs in different chromosomes were identified. As these studies have used different sets of markers and many of the QTLs identified are long, it is difficult to exploit the varietal difference for improved root traits by marker assisted selection and for identification of concerned alleles. Intensive data mining of literature resulted in the identification 861 root development QTLs and associated microsatellite markers located on different chromosomes. The QTL and marker data generated and the genome sequence of rice were used for construction of a relational database, Rootbrowse, using MySQL relational database management system and Bio::DB::GFF schema. The data is viewed using GBrowse visualization tool. It graphically displays a section of the genome and all features annotated on it including the QTLs. The QTLs can be displayed along with SSR markers, protein coding genes and/or known root development genes for prediction of probable candidate genes.     

Inhibition of matrix metalloproteinase-2 by PARP inhibitors

Matrix metalloproteinase-2 (MMP-2), a ubiquitously expressed zinc-dependent endopeptidase, and poly(ADP-ribosyl) polymerase (PARP), a nuclear enzyme regulating DNA repair, are activated by nitroxidative stress associated with various pathologies. As MMP-2 plays a detrimental role in heart injuries resulting from enhanced nitroxidative stress, where PARP and MMP inhibitors are beneficial, we hypothesized that PARP inhibitors may affect MMP-2 activity. Using substrate degradation assays to determine MMP-2 activity we found that four PARP inhibitors (3-AB, PJ-34, 5-AIQ, and EB-47) inhibited 64 kDa MMP-2 in a concentration-dependent manner. The IC₅₀ values of PJ-34 and 5-AIQ were in the high micromolar range and comparable to those of known MMP-2 inhibitors doxycycline, minocycline or o-phenanthroline, whereas those for 3-AB and EB-47 were in the millimolar range. Co-incubation of PARP inhibitors with doxycycline showed an additive inhibition of MMP-2 that was significant for 3-AB alone. These data demonstrate that the protective effects of some PARP inhibitors may include inhibition of MMP-2 activity.     

Occasional intraguild predation structuring small mammal assemblages: the marsupial Didelphis aurita in the Atlantic Forest of Brazil

The didelphid marsupial, Didelphis aurita, is suggested as an intraguild predator and as key‐species in small mammal assemblages of the Atlantic Forest of Brazil. The field experiments required to test this hypothesis are complex to implement, but the recent revival of regression methods offers a viable alternative. Here we use the dynamic and static regression methods to determine the importance of D. aurita as a competitor and intraguild predator. Capture–recapture data from two localities in the Rio de Janeiro State were used, Garrafão (municipality of Guapimirim), a coastal forest of the Serra do Mar, and Barra de Maricá, a costal sand dune vegetation. Population and microhabitat variables were monitored from April 1997 to April 2003 in Garrafão, and from January 1986 to July 1990 in Barra de Maricá. Microhabitat variables were related to Canopy, Plant, Litter and Rock covers, Obstruction from 0 to 1.5 m, and Number of logs. Exploitation competition was tested by the dynamic method, which models the effects of D. aurita on the per capita growth rate of a species. Interference by predation or competition was tested by the static method, where the abundance of D. aurita at trap stations was regressed against the abundance of other small mammals, after removal of any variation associated with microhabitat factors. Exploitation competition was not detected, but the interference of D. aurita was pervasive, affecting all small mammals studied in the two localities. The clear avoidance of D. aurita by all small mammals tested in two localities of different physiognomies indicates that it functions as an intraguild predator, even if actual predation by D. aurita is an occasional event.     

Immobilization of acidic lipase derived from Pseudomonas gessardii onto mesoporous activated carbon for the hydrolysis of olive oil

Mesoporous activated carbon (MAC) derived from rice husk is used for the immobilization of acidic lipase (ALIP) produced from Pseudomonas gessardii. The purified acidic lipase had the specific activity and molecular weight of 1473 U/mg and 94 kDa respectively. To determine the optimum conditions for the immobilization of lipase onto MAC, the experiments were carried out by varying the time (10–180 min), pH (2–8), temperature (10–50 °C) and the initial lipase activity (49 × 10³, 98 × 10³, 147 × 10³ and 196 × 10³ U/l in acetate buffer). The optimum conditions for immobilization of acidic lipase were found to be: time—120 min; pH 3.5; temperature—30 °C, which resulted in achieving a maximum immobilization of 1834 U/g. The thermal stability of the immobilized lipase was comparatively higher than that in its free form. The free and immobilized enzyme kinetic parameters (K_m and V_max) were found using Michaelis–Menten enzyme kinetics. The K_m values for free enzyme and immobilized one were 0.655 and 0.243 mM respectively. The immobilization of acidic lipase onto MAC was confirmed using Fourier Transform-Infrared Spectroscopy, X-ray diffraction analysis and scanning electron microscopy.     

Formulation and Optimization of Zidovudine Niosomes

Zidovudine (AZT) is commonly used to treat patients with AIDS, but it is limited by toxicity and high dosing needs. Alternative formulations have been proposed to overcome these drawbacks. The objective of this study was to evaluate process-related variables like hydration and sonication time, rotation speed of evaporation flask, and the effects of charge-inducing agent and centrifugation on zidovudine entrapment and release from niosomes. Formulation of zidovudine niosomes was optimized by altering the proportions of Tween, Span and cholesterol. The effect of process–related variables like hydration time, sonication time, charge-inducing agent, centrifugation and rotational speed of evaporation flask on zidovudine entrapment and release from niosomes was evaluated. The effect of changes in osmotic shock and viscosity were also evaluated. Non-sonicated niosomes were in the size range of 2-3.5 μm and sonicated niosomes formulated with Tween 80 and dicetylphosphate (DCP) had a mean diameter of 801 nm. Zidovudine niosomes formulated with Tween 80 entrapped high amounts of drug and the addition of DCP enhanced drug release for a longer time (88.72% over 12 h). The mechanism of release from Tween 80 formulation was the Fickian type and obeyed first-order release kinetics. Niosomes can be formulated by proper adjustment of process parameters to enhance zidovudine entrapment and sustainability of release. These improvements in zidovudine formulation may be useful in developing a more effective AIDS therapy.     

toxin-mediated insect resistance in plants

We are currently in an interesting phase of plant biotechnology releases, both for the scientists responsible for these innovations who are beginning to see their ideas realized, and for the biotechnology companies that are starting to see a return on their investment. One of the most notable examples, is the introduction of transgenic crops that are engineered to express a Bacillus thuringiensis toxin that confers resistance to insect predation. However, the picture is not altogether positive - there is concern that the introduction of this technology was premature or should not have happened at all, and that the valuable insecticidal properties of Bacillus thuringiensis will be lost.