Identification and characterization of tac5, a telomerase activation mutant, characterization of DNA damage responses and assessment of interactions between telomere-related proteins in Arabidopsis thaliana

statistics, 2) unadjusted inferential statistics, 3) stratified analysis, and 4) multivariable models. My investigation produced results in accord with generally accepted notions in addition to significant findings that interestingly counter current preconceptions. Intraspecies contact was more common than inter-species, with indirect contact occurring more frequently than direct. Direct contact between species occurred extremely rarely. The most important factors that influenced the rate of contact for both species were water, winter, and cultivated fields. Information regarding probability of infectious agent survival and transfer will be used in the future to advance current epidemiological models, including geographicautomata (Ward et al. 2007: In Press) and cellular automata models (Doran and Laffan 2005) to better understand and manage integrated domestic cattle and free-ranging wildlife populations. Such modeling provides essential and necessary knowledge for developing prevention, detection, response, and recovery strategies – employed in advance, during, and after a disease outbreak, respectively. responsible for telomere activation. In addition tac5 showed sensitivity to hydrogen peroxide treatment, suggesting a novel role of telomerase in the mitochondrial environment. Chapter III reports the role of PARP proteins in plant telomere biology. Both AtPARP1 and AtPARP2 are transcriptionally upregulated in response to DNA damage treatment or telomere dysfunction. However, in contrast to mammalian PARPs, the Arabidopsis proteins do not appear to have a function in telomere length maintenance as indicated by TRF analysis or in promoting genome stability maintenance as indicated by cytogenetic studies. Further analysis of PARP interactions at dysfunctional telomeres in the genetically tractable Arabidopsis model may provide insight into the cellular response to dysfunctional telomeres. As explained in chapter IV, the yeast two-hybrid screen was utilized to confirm the interactions of ATR with AtPOT2 and Ku80 and to identify novel interacting partners of Arabidopsis telomere proteins. At2g04410 (Unknown protein) was identified as a direct interacting partner of AtPOT1. This interaction was confirmed in vitro by coimmunoprecipitation assay. Further analysis of the unknown protein may shed light on AtPOT1’s function in telomere maintenance

Use of polyethyleneimine polymer in cell culture as attachment factor and lipofection enhancer

BACKGROUND: Several cell lines and primary cultures benefit from the use of positively charged extracellular matrix proteins or polymers that enhance their ability to attach to culture plates. Polyethyleneimine is a positively charged polymer that has gained recent attention as a transfection reagent. A less known use of this cationic polymer as an attachment factor was explored with several cell lines. RESULTS: Polyethyleneimine compared favorably to traditional attachment factors such as collagen and polylysine. PC-12 and HEK-293 cells plated on dishes coated with polyethyleneimine showed a homogeneous distribution of cells in the plate, demonstrating strong cell adhesion that survived washing procedures. The polymer could also be used to enhance the adherence and allow axonal outgrowth from zebrafish retinal explants. The effects of this coating agent on the transfection of loosely attaching cell lines were studied. Pre-coating with polyethyleneimine had the effect of enhancing the transfection yield in procedures using lipofection reagents. CONCLUSION: Polyethyleneimine is an effective attachment factor for weakly anchoring cell lines and primary cells. Its use in lipofection protocols makes the procedures more reliable and increases the yield of expressed products with commonly used cell lines such as PC-12 and HEK-293 cells

Genome-wide histone state profiling of fibroblasts from the opossum, Monodelphis domestica, identifies the first marsupial-specific imprinted gene

BACKGROUND: Imprinted genes have been extensively documented in eutherian mammals and found to exhibit significant interspecific variation in the suites of genes that are imprinted and in their regulation between tissues and developmental stages. Much less is known about imprinted loci in metatherian (marsupial) mammals, wherein studies have been limited to a small number of genes previously known to be imprinted in eutherians. We describe the first ab initio search for imprinted marsupial genes, in fibroblasts from the opossum, Monodelphis domestica, based on a genome-wide ChIP-seq strategy to identify promoters that are simultaneously marked by mutually exclusive, transcriptionally opposing histone modifications. RESULTS: We identified a novel imprinted gene (Meis1) and two additional monoallelically expressed genes, one of which (Cstb) showed allele-specific, but non-imprinted expression. Imprinted vs. allele-specific expression could not be resolved for the third monoallelically expressed gene (Rpl17). Transcriptionally opposing histone modifications H3K4me3, H3K9Ac, and H3K9me3 were found at the promoters of all three genes, but differential DNA methylation was not detected at CpG islands at any of these promoters. CONCLUSIONS: In generating the first genome-wide histone modification profiles for a marsupial, we identified the first gene that is imprinted in a marsupial but not in eutherian mammals. This outcome demonstrates the practicality of an ab initio discovery strategy and implicates histone modification, but not differential DNA methylation, as a conserved mechanism for marking imprinted genes in all therian mammals. Our findings suggest that marsupials use multiple epigenetic mechanisms for imprinting and support the concept that lineage-specific selective forces can produce sets of imprinted genes that differ between metatherian and eutherian lines

Transfection yields were determined by β-galactosidase assays and normalized to the yields obtained with PEI coating of the culture dish (to which it was assigned an arbitrary value of 100

<p><b>Copyright information:</b></p><p>Taken from "Use of polyethyleneimine polymer in cell culture as attachment factor and lipofection enhancer"</p><p>BMC Biotechnology 2004;4():23-23.</p><p>Published online 15 Oct 2004</p><p>PMCID:PMC526208.</p><p>Copyright © 2004 Vancha et al; licensee BioMed Central Ltd.</p>0%). PEI and other attachment factors enhanced the transfection of loosely attaching cell lines such as PC-12 cells (graph A) and HEK-293 cells (graph B). No significant effect was observed with tightly attaching cells such as MYS fibroblasts (graph C). The bars show the average ± standard deviation of triplicate assays, which are representative of at least two independent experiments (* significantly higher than untreated control, p < 0.01)

Permanent Genetic Resources added to Molecular Ecology Resources Database 1 February 2011–31 March 2011

This article documents the addition of 111 microsatellite marker loci to the Molecular Ecology Resources Database. Loci were developed for the following species: Acipenser oxyrinchus desotoi, Anopheles nuneztovari sensu lato, Asellus aquaticus, Calopteryx splendens, Calopteryx virgo, Centaurea aspera, Centaurea seridis, Chilina dombeyana, Proctoeces cf. lintoni and Pyrenophora teres f. teres

Localization of genes for V+LDL plasma cholesterol levels on two diets in the opossum Monodelphis domestica[S]

Plasma cholesterol levels among individuals vary considerably in response to diet. However, the genes that influence this response are largely unknown. Non-HDL (V+LDL) cholesterol levels vary dramatically among gray, short-tailed opossums fed an atherogenic diet, and we previously reported that two quantitative trait loci (QTLs) influenced V+LDL cholesterol on two diets. We used hypothesis-free, genome-wide linkage analyses on data from 325 pedigreed opossums and located one QTL for V+LDL cholesterol on the basal diet on opossum chromosome 1q [logarithm of the odds (LOD) = 3.11, genomic P = 0.019] and another QTL for V+LDL on the atherogenic diet (i.e., high levels of cholesterol and fat) on chromosome 8 (LOD = 9.88, genomic P = 5 × 10−9). We then employed a novel strategy involving combined analyses of genomic resources, expression analysis, sequencing, and genotyping to identify candidate genes for the chromosome 8 QTL. A polymorphism in ABCB4 was strongly associated (P = 9 × 10−14) with the plasma V+LDL cholesterol concentrations on the high-cholesterol, high-fat diet. The results of this study indicate that genetic variation in ABCB4, or closely linked genes, is responsible for the dramatic differences among opossums in their V+LDL cholesterol response to an atherogenic diet