Atomic Force Microscope nanolithography on chromosomes to genrate single-cell genetic probes

Abstract Background Chromosomal dissection provides a direct advance for isolating DNA from cytogenetically recognizable region to generate genetic probes for fluorescence in situ hybridization, a technique that became very common in cyto and molecular genetics research and diagnostics. Several reports describing microdissection methods (glass needle or a laser beam) to obtain specific probes from metaphase chromosomes are available. Several limitations are imposed by the traditional methods of dissection as the need for a large number of chromosomes for the production of a probe. In addition, the conventional methods are not suitable for single chromosome analysis, because of the relatively big size of the microneedles. Consequently new dissection techniques are essential for advanced research on chromosomes at the nanoscale level. Results We report the use of Atomic Force Microscope (AFM) as a tool for nanomanipulation of single chromosomes to generate individual cell specific genetic probes. Besides new methods towards a better nanodissection, this work is focused on the combination of molecular and nanomanipulation techniques which enable both nanodissection and amplification of chromosomal and chromatidic DNA. Cross-sectional analysis of the dissected chromosomes reveals 20 nm and 40 nm deep cuts. Isolated single chromosomal regions can be directly amplified and labeled by the Degenerate Oligonucleotide-Primed Polymerase Chain Reaction (DOP-PCR) and subsequently hybridized to chromosomes and interphasic nuclei. Conclusions Atomic force microscope can be easily used to visualize and to manipulate biological material with high resolution and accuracy. The fluorescence in situ hybridization (FISH) performed with the DOP-PCR products as test probes has been tested succesfully in avian microchromosomes and interphasic nuclei. Chromosome nanolithography, with a resolution beyond the resolution limit of light microscopy, could be useful to the construction of chromosome band libraries and to the molecular cytogenetic mapping related to the investigation of genetic diseases.</p

Novel atrazine-binding biomimetics inspired to the D1 protein from the photosystem II of Chlamydomonas reinhardtii

Biomimetic design represents an emerging field for improving knowledge of natural molecules, as well as to project novel artificial tools with specific functions for biosensing. Effective strategies have been exploited to design artificial bioreceptors, taking inspiration from complex supramolecular assemblies. Among them, size-minimization strategy sounds promising to provide bioreceptors with tuned sensitivity, stability, and selectivity, through the ad hoc manipulation of chemical species at the molecular scale. Herein, a novel biomimetic peptide enabling herbicide binding was designed bioinspired to the D1 protein of the Photosystem II of the green alga Chlamydomonas reinhardtii. The D1 protein portion corresponding to the QB plastoquinone binding niche is capable of interacting with photosynthetic herbicides. A 50-mer peptide in the region of D1 protein from the residue 211 to 280 was designed in silico, and molecular dynamic simulations were performed alone and in complex with atrazine. An equilibrated structure was obtained with a stable pocked for atrazine binding by three H-bonds with SER222, ASN247, and HIS272 residues. Computational data were confirmed by fluorescence spectroscopy and circular dichroism on the peptide obtained by automated synthesis. Atrazine binding at nanomolar concentrations was followed by fluorescence spectroscopy, highlighting peptide suitability for optical sensing of herbicides at safety limits

Rapporto sull’attività 29 Giugno – 6 Luglio 2013

In questo lavoro viene descritta la campagna geofisica e idro-oceanografica ‘GARGANO2013’ effettuata grazie ad una collaborazione tra INGV e Marina Militare Italiana nell’ambito di un accordo di collaborazione denominato CONAGEM (Coordinamento Nazionale per la Geofisica Marina). Tale accordo, siglato nel 2005 fra i principali enti pubblici di ricerca che operano in mare, prevede infatti la possibilità di attuare congiuntamente campagne di ricerca marine condividendone dati e risultati. La campagna è stata organizzata con la finalità di raccogliere quanti più possibili elementi utili a caratterizzare l’area marina del Gargano sotto un profilo ambientale predisponendone un quadro di riferimento per successive attività di monitoraggio delle fenomenologie presenti nell’area di indagine. In quest’ottica, l’obiettivo prefissato era la mappatura di eventuali affioramenti di acque dolci in mare. Contestualmente, è stata eseguita la sperimentazione di prototipi strumentali per future applicazioni a bordo nave: in particolare, sono state eseguite misure di tipo accelerometrico al fine di caratterizzare le sollecitazioni dinamiche cui sono sottoposti gli strumenti a bordonave ottenendo così utili informazioni per lo sviluppo di nuove tecnologie quali piattaforme inerziali per uso scientifico e strumentazione per prospezioni gravimetriche da utilizzare su nave

Carbonaceous aerosol in polar areas: First results and improvements of the sampling strategies

While more and more studies are being conducted on carbonaceous fractions—organic carbon (OC) and elemental carbon (EC)—in urban areas, there are still too few studies about these species and their effects in polar areas due to their very low concentrations; further, studies in the literature report only data from intensive campaigns, limited in time. We present here for the first time EC–OC concentration long-time data records from the sea-level sampling site of Ny-Ålesund, in the High Arctic (5 years), and from Dome C, in the East Antarctic Plateau (1 year). Regarding the Arctic, the median (and the interquartile range (IQR)) mass concentrations for the years 2011–2015 are 352 (IQR: 283–475) ng/m3 for OC and 4.8 (IQR: 4.6–17.4) ng/m3 for EC, which is responsible for only 3% of total carbon (TC). From both the concentration data sets and the variation of the average monthly concentrations, the influence of the Arctic haze on EC and OC concentrations is evident. Summer may be interested by high concentration episodes mainly due to long-range transport (e.g., from wide wildfires in the Northern Hemisphere, as happened in 2015). The average ratio of EC/OC for the summer period is 0.05, ranging from 0.02 to 0.10, and indicates a clean environment with prevailing biogenic (or biomass burning) sources, as well as aged, highly oxidized aerosol from long-range transport. Contribution from ship emission is not evident, but this result may be due to the sampling time resolution. In Antarctica, a 1 year-around data set from December 2016 to February 2018 is shown, which does not present a clear seasonal trend. The OC median (and IQR) value is 78 (64–106) ng/m3; for EC, it is 0.9 (0.6–2.4) ng/m3, weighing for 3% on TC values. The EC/OC ratio mean value is 0.20, with a range of 0.06–0.35. Due to the low EC and OC concentrations in polar areas, correction for the blank is far more important than in campaigns carried out in other regions, largely affecting uncertainties in measured concentrations. Through the years, we have thus developed a new sampling strategy that is presented here for the first time: samplers were modified in order to collect a larger amount of particulates on a small surface, enhancing the capability of the analytical method since the thermo-optical analyzer is sensitive to carbonaceous aerosol areal density. Further, we have recently coupled such modified samplers with a sampling strategy that makes a more reliable blank correction of every single sample possible

LABEC, the INFN ion beam laboratory of nuclear techniques for environment and cultural heritage

The LABEC laboratory, the INFN ion beam laboratory of nuclear techniques for environment and cultural heritage, located in the Scientific and Technological Campus of the University of Florence in Sesto Fiorentino, started its operational activities in 2004, after INFN decided in 2001 to provide our applied nuclear physics group with a large laboratory dedicated to applications of accelerator-related analytical techniques, based on a new 3 MV Tandetron accelerator. The new accelerator greatly improved the performance of existing Ion Beam Analysis (IBA) applications (for which we were using since the 1980s an old single-ended Van de Graaff accelerator) and in addition allowed to start a novel activity of Accelerator Mass Spectrometry (AMS), in particular for 14C dating. Switching between IBA and AMS operation became very easy and fast, which allowed us high flexibility in programming the activities, mainly focused on studies of cultural heritage and atmospheric aerosol composition, but including also applications to biology, geology, material science and forensics, ion implantation, tests of radiation damage to components, detector performance tests and low-energy nuclear physics. This paper describes the facilities presently available in the LABEC laboratory, their technical features and some success stories of recent applications

Rapporto sull’attività 29 Giugno – 6 Luglio 2013

In questo lavoro viene descritta la campagna geofisica e idro-oceanografica ‘GARGANO2013’ effettuata grazie ad una collaborazione tra INGV e Marina Militare Italiana nell’ambito di un accordo di collaborazione denominato CONAGEM (Coordinamento Nazionale per la Geofisica Marina). Tale accordo, siglato nel 2005 fra i principali enti pubblici di ricerca che operano in mare, prevede infatti la possibilità di attuare congiuntamente campagne di ricerca marine condividendone dati e risultati. La campagna è stata organizzata con la finalità di raccogliere quanti più possibili elementi utili a caratterizzare l’area marina del Gargano sotto un profilo ambientale predisponendone un quadro di riferimento per successive attività di monitoraggio delle fenomenologie presenti nell’area di indagine. In quest’ottica, l’obiettivo prefissato era la mappatura di eventuali affioramenti di acque dolci in mare. Contestualmente, è stata eseguita la sperimentazione di prototipi strumentali per future applicazioni a bordo nave: in particolare, sono state eseguite misure di tipo accelerometrico al fine di caratterizzare le sollecitazioni dinamiche cui sono sottoposti gli strumenti a bordonave ottenendo così utili informazioni per lo sviluppo di nuove tecnologie quali piattaforme inerziali per uso scientifico e strumentazione per prospezioni gravimetriche da utilizzare su nave.Marina Militare ItalianaPublished1-283A. Ambiente MarinoN/A or not JCRope

Sea salt sodium record from Talos Dome (East Antarctica) as a potential proxy of the Antarctic past sea ice extent

Antarctic sea ice has shown an increasing trend in recent decades, but with strong regional differences from one sector to another of the Southern Ocean. The Ross Sea and the Indian sectors have seen an increase in sea ice during the satellite era (1979 onwards). Here we present a record of ssNa+ flux in the Talos Dome region during a 25-year period spanning from 1979 to 2003, showing that this marker could be used as a potential proxy for reconstructing the sea ice extent in the Ross Sea and Western Pacific Ocean at least for recent decades. After finding a positive relationship between the maxima in sea ice extent for a 25-year period, we used this relationship in the TALDICE record in order to reconstruct the sea ice conditions over the 20th century. Our tentative reconstruction highlighted a decline in the sea ice extent (SIE) starting in the 1950s and pointed out a higher variability of SIE starting from the 1960s and that the largest sea ice extents of the last century occurred during the 1990s

Structural and Functional Hierarchy in Photosynthetic Energy Conversion—from Molecules to Nanostructures

Basic principles of structural and functional requirements of photosynthetic energy conversion in hierarchically organized machineries are reviewed. Blueprints of photosynthesis, the energetic basis of virtually all life on Earth, can serve the basis for constructing artificial light energy-converting molecular devices. In photosynthetic organisms, the conversion of light energy into chemical energy takes places in highly organized fine-tunable systems with structural and functional hierarchy. The incident photons are absorbed by light-harvesting complexes, which funnel the excitation energy into reaction centre (RC) protein complexes containing redox-active chlorophyll molecules; the primary charge separations in the RCs are followed by vectorial transport of charges (electrons and protons) in the photosynthetic membrane. RCs possess properties that make their use in solar energy-converting and integrated optoelectronic systems feasible. Therefore, there is a large interest in many laboratories and in the industry toward their use in molecular devices. RCs have been bound to different carrier matrices, with their photophysical and photochemical activities largely retained in the nano-systems and with electronic connection to conducting surfaces. We show examples of RCs bound to carbon-based materials (functionalized and non-functionalized single- and multiwalled carbon nanotubes), transitional metal oxides (ITO) and conducting polymers and porous silicon and characterize their photochemical activities. Recently, we adapted several physical and chemical methods for binding RCs to different nanomaterials. It is generally found that the P(+)(Q(A)Q(B))(−) charge pair, which is formed after single saturating light excitation is stabilized after the attachment of the RCs to the nanostructures, which is followed by slow reorganization of the protein structure. Measuring the electric conductivity in a direct contact mode or in electrochemical cell indicates that there is an electronic interaction between the protein and the inorganic carrier matrices. This can be a basis of sensing element of bio-hybrid device for biosensor and/or optoelectronic applications