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Frozen bacterial cells were low-temperature embedded after cryosubstitution at 185K in organic solvent. Temperature elevation of cryosubstituted E. coli cells, when still in organic solvent, had no effect on the preservation of chromatin structure. The achieved stabilization was found to be established independent of the presence of the chemical fixative. Cryosubstitution without the use of a chemical...
In this essay with a flavor of science history, the influence of imaging techniques, as compared to other research strategies, on microbiologic investigations is discussed. Using a few selected examples, to what degree microbiology became a leading science during the last 50 years in gaining knowledge in life sciences, particularly with regard to molecular genetics and more recently molecular evolution...
Various DNA-based methods are presently being applied for identification of industrial bacterial cultures including dairy starter and probiotic strains of Lactobacillus. The success of strain-specific identification depends on the power of the DNA-based methods to reveal intraspecies DNA polymorphism. This study reveals that all eleven arbitrarily chosen Lactobacillus rhamnosus starter, laboratory...
The goal of this study was to construct high resolution 3D confocal images of regions of condensed and extended chromatin in cell nuclei and individual chromosomes. It has been shown previously that sensitivity of DNA in situ to denaturation correlates with chromatin condensation and varies during cell cycle and apoptosis. Thus, detection of DNA which was partially denatured in situ provided a means...
The interactions of the several linear DNA fragments and cisplatin at different concentrations and incubation time have been studied by atomic force microscopy. When equal volume of 2mM cisplatin and 5ng/μl 1134bp DNA fragment were incubated for 30min, 1h, 2h and 3h, the observed DNA were gradually changed from the extended chains to the bending structures, the worm-like structures and flat particles...
Sample preparation techniques allowing reliable and reproducible imaging of DNA with various structures, topologies and complexes with proteins are reviewed. The major emphasis is given to methods utilizing chemical functionalization of mica, enabling preparation of the surfaces with required characteristics. The methods are illustrated by examples of imaging of different DNA structures. Special attention...
This paper examines two fabrication techniques (soft lithography and UV embossing) employed in the replication of various structures at the micron and single nanometer regimes. Stretched and assembled forms of double stranded (ds) DNA (16–3μm in length) were adsorbed on coated silicon oxide. The collective results show that the resolution of all the thermally cured polymers improves significantly...
Heterochromatin bodies in single- and multichromocentered interphase cell nuclei of Triatoma infestans, a vector of Chagas disease, have been suggested to contain AT-rich DNA, based on their positive response to Q-banding and Hoechst 33248 treatment. No information exists on whether GC-rich DNA is also present in these nuclei and whether it plays a role on chromatin condensation. Considering that...
Transcription factors play a central role in cell biology through binding to target DNA elements and regulating gene expression. In this study, we used the p53 tumour suppressor as a model transcription factor to develop an imaging based assay to measure DNA binding. The assay utilizes fluorescence imaging microscopy to detect labelled p53 bound to DNA coated on microbeads. We demonstrate the ability...
The purpose of this review is to discuss the achievements and progress that has been made in the use of atomic force microscopy in DNA related research in the last 25 years. For this review DNA related research is split up in chromosomal-, chromatin- and DNA focused research to achieve a logical flow from large- to smaller structures. The focus of this review is not only on the AFM as imaging tool...
The Fourier transform-infrared (FT-IR) signature of dry samples of DNA and DNA-polypeptide complexes, as studied by IR microspectroscopy using a diamond attenuated total reflection (ATR) objective, has revealed important discriminatory characteristics relative to the PO 2− vibrational stretchings. However, DNA IR marks that provide information on the sample's richness in hydrogen bonds...
Oxaliplatin is one of the most important anticancer drugs at present. However, the mechanism of action of oxaliplatin is still controversial. In this study, the interactions between oxaliplatin and a plasmid DNA have been studied so as to reveal the structural basis of its activity. The structural characteristic of pUC19 DNA (2ng/μL) incubated with 100μmol/L and 1000μmol/L of oxaliplatin for the different...
Nucleic acids are responsible for the storage, transfer and realization of genetic information in the cell, which provides correct development and functioning of organisms. DNA interaction with ligands ensures the safety of this information. Over the past 10 years, advances in electron microscopy and image processing allowed to obtain the structures of key DNA-protein complexes with resolution below...
While the widespread emergence of nanoscience and nanotechnology can be dated back to the early eighties, the last decade has witnessed a true coming of age of this research field, with novel nanomaterials constantly finding their way into marketed products. The performance of nanomaterials being dominated by their nanoscale morphology, their quantitative characterization with respect to a number...
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