Search results for "Fluorescence"
showing 10 items of 2463 documents
Asynchronous replication dynamics of imprinted and non-imprinted chromosome regions in early mouse embryos.
2008
We have used interphase FISH to analyze the replication behavior of four imprinted chromosome regions (Snrpn, Zim1-Peg3, Dlk1-Gtl2, and Igf2r) and five non-imprinted regions in mouse one-cell to morula-stage embryos and embryonic fibroblasts. In general, imprinted chromosome regions showed the expected asynchronous pattern of replication throughout all analyzed stages of preimplantation development and in differentiated cells. The Dlk1-Gtl2 locus which is not expressed and Igf2r which is biallelically expressed in early embryos showed a relaxation of replication asynchrony at the morula stage. Asynchronous replication in zygotes and two-cell embryos was not specific to imprinted regions. Th…
Processing of O6-methylguanine into DNA double-strand breaks requires two rounds of replication whereas apoptosis is also induced in subsequent cell …
2009
The DNA adduct O(6)-methylguanine (O(6)MeG) induced by environmental genotoxins and anticancer drugs is a highly mutagenic, genotoxic and apoptotic lesion. Apoptosis induced by O(6)MeG requires mismatch repair (MMR) and proliferation. Models of O(6)MeG-triggered cell death postulate that O(6)MeG/T mispairs activate MMR giving rise to either direct genotoxic signaling or secondary lesions that trigger apoptotic signaling in the 2(nd) replication cycle. To test these hypotheses, we used a highly synchronized cell system competent and deficient for the repair of O(6)MeG adducts, which were induced by the S(N)1 methylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). We show that DNA doub…
The Translesion Polymerase Rev3L in the Tolerance of Alkylating Anticancer Drugs
2009
Temozolomide and fotemustine, representing methylating and chloroethylating agents, respectively, are used in the treatment of glioma and malignant melanoma. Because chemoresistance of these tumors is a common phenomenon, identification of the underlying mechanisms is needed. Here we show that Rev3L, the catalytic subunit of the translesion DNA polymerase zeta, mediates resistance to both temozolomide and fotemustine. Rev3L knockout cells are hypersensitive to both agents. It is remarkable that cells heterozygous for Rev3L showed an intermediate sensitivity. Rev3L is not involved in the tolerance of the toxic O6-methylguanine lesion. However, a possible role of Rev3L in the tolerance of O6-…
On the Effect of Downscaling in Inkjet Printed Life-Inspired Compartments
2019
The fabrication of size-scalable liquid compartments is a topic of fundamental importance in synthetic biology, aiming to mimic the structures and the functions of cellular compartments. Here, inkjet printing is demonstrated as a customizable approach to fabricate aqueous compartments at different size regimes (from nanoliter to femtoliter scale) revealing the crucial role of size in governing the emerging of new properties. At first, inkjet printing is shown to produce homogenous aqueous compartments stabilized by oil-confinement with mild surfactants down to the hundreds of picoliter scale [1]. Raster Image Correlation Spectroscopy allows to monitor few intermolecular events by the involv…
CY5 fluorescence measured with silicon photomultipliers
2014
This paper presents an efficient optical biosensor set up for a low-level light detection, using fluorescent dyes and a novel Si-based detector. Fluorescence emitted by a traditional fluorophore, CY5, widely used as optical label in DNA microarrays, was detected using a 25 pixels Silicon photomultiplier (SiPM), a device formed by avalanche diodes operating in Geiger mode, in parallel connections. We measured the fluorescence current in different deposition (fluorophore concentration; solvent; salt concentration) and operation (angle of analysis, optical laser power, device gain) conditions. The characterization of DNA samples labeled with CY5 is also reported to demonstrate the detector pot…
Investigating bacterial populations in styrene-degrading biofilters by 16S rDNA tag pyrosequencing
2014
Microbial biofilms are essential components in the elimination of pollutants within biofilters, yet still little is known regarding the complex relationships between microbial community structure and biodegradation function within these engineered ecosystems. To further explore this relationship, 16S rDNA tag pyrosequencing was applied to samples taken at four time points from a styrene-degrading biofilter undergoing variable operating conditions. Changes in microbial structure were observed between different stages of biofilter operation, and the level of styrene concentration was revealed to be a critical factor affecting these changes. Bacterial genera Azoarcus and Pseudomonas were among…
A Sensitive Method for Identification of DNA Dependent DNA Polymerases in Acrylamide Gels after Seperation by Micro Disc Electrophoresis
1973
Abstract DNA polymerase, disc electrophoresis, template affinity Two sensitive methods are described for detection of DNA dependent DNA polymerase activities in polyacrylamide gels after their fractionation by micro-disc electrophoresis. One technique is based on the increase in fluorescence of the ethidium bromide complex with template polydeoxyribonucleotides brought about by the action of the polymerases. The sensitivity of the previously described technique has been enhanced. Another method, 14 fold as sensitive, uses radioactive precursors in the enzyme assay after electrophoretic separation; washing, slicing and counting allows to evaluate incorporation into acid insoluble polymer, re…
Distribution and diversity of type III secretion system-like genes in saprophytic and phytopathogenic fluorescent pseudomonads
2004
Type three secretion systems (TTSSs) are protein translocation mechanisms associated with bacterial pathogenicity in host plants, and hypersensitive reactions in non-host plants. Distribution and diversity of TTSS-like genes within a collection of saprophytic and phytopathogenic fluorescent pseudomonads were characterized. This collection included 16 strains belonging to 13 pathogenic species, and 87 strains belonging to five saprophytic species isolated from plant rhizosphere and soil. Presence of conserved hypersensitive reaction/pathogenicity (hrp) genes (hrc RST) was assessed both by PCR using primers designed to amplify the corresponding sequence and by dot-blot hybridization using a P…
Fast protocols for the 5S rDNA and ITS-2 based identification ofOenococcus oeni
2005
To identify specific marker sequences for the rapid identification of Oenococcus oeni, we sequenced the 23S-5S internal transcribed spacer (ITS-2) region and the 5S rDNA of five different O. oeni strains and three phylogenetically related lactic acid bacteria (LAB). Comparative analysis revealed 100% identity among the ITS-2 region of the O. oeni strains and remarkable differences in length and sequence compared to related LAB. These results enabled us to develop a primer set for a rapid PCR-identification of O. oeni within three hours. Moreover, the comparison of the 5S rDNA sequences and the highly conserved secondary structure provided the template for the design of three fluorescence-la…
Isolation of methanotrophic bacteria from termite gut.
2015
The guts of termites feature suitable conditions for methane oxidizing bacteria (MOB) with their permanent production of CH4 and constant supply of O2 via tracheae. In this study, we have isolated MOB from the gut contents of the termites Incisitermes marginipennis, Mastotermes darwiniensis, and Neotermes castaneus for the first time. The existence of MOB was indicated by detecting pmoA, the gene for the particulate methane monooxygenase, in the DNA of gut contents. Fluorescence in situ hybridization and quantitative real-time polymerase chain reaction supported those findings. The MOB cell titer was determined to be 10(2)-10(3) per gut. Analyses of the 16S rDNA from isolates indicated clos…