Non-topographic current contrast in scanning field emission microscopy.

In scanning field emission microscopy (SFEM), a tip (the source) is approached to few (or a few tens of) nanometres distance from a surface (the collector) and biased to field-emit electrons. In a previous study (Zanin et al. 2016 Proc. R. Soc. A 472, 20160475. (doi:10.1098/rspa.2016.0475)), the field-emitted current was found to change by approximately 1% at a monatomic surface step (approx. 200 pm thick). Here we prepare surface domains of adjacent different materials that, in some instances, have a topographic contrast smaller than 15 pm. Nevertheless, we observe a contrast in the field-emitted current as high as 10%. This non-topographic collector material dependence is a yet unexplored degree of freedom calling for a new understanding of the quantum mechanical tunnelling barrier at the source site that takes into account the properties of the material at the collector site.

Experimental phase diagram of perpendicularly magnetized ultrathin ferromagnetic films.

We image the domain patterns in perpendicularly magnetized ultrathin Fe films on Cu(100) as a function of the temperature T and the applied magnetic field H. Between the low-field stripe phase and the high-field uniform phase we find a bubble phase, consisting of reversed circular domains in a homogeneous background. The curvature of the transition lines in the H-T parameter space is in contrast to the general expectations. The pattern transformations show yet undetected scaling properties.

The composite genome of the legume symbiont Sinorhizobium meliloti.

The scarcity of usable nitrogen frequently limits plant growth. A tight metabolic association with rhizobial bacteria allows legumes to obtain nitrogen compounds by bacterial reduction of dinitrogen (N2) to ammonium (NH4+). We present here the annotated DNA sequence of the alpha-proteobacterium Sinorhizobium meliloti, the symbiont of alfalfa. The tripartite 6.7-megabase (Mb) genome comprises a 3.65-Mb chromosome, and 1.35-Mb pSymA and 1.68-Mb pSymB megaplasmids. Genome sequence analysis indicates that all three elements contribute, in varying degrees, to symbiosis and reveals how this genome may have emerged during evolution. The genome sequence will be useful in understanding the dynamics of interkingdom associations and of life in soil environments.

Electron beam confinement and image contrast enhancement in near field emission scanning electron microscopy.

In conventional scanning electron microscopy (SEM), the lateral resolution is limited by the electron beam diameter impinging on the specimen surface. Near field emission scanning electron microscopy (NFESEM) provides a simple means of overcoming this limit; however, the most suitable field emitter remains to be determined. NFESEM has been used in this work to investigate the W (110) surface with single-crystal tungsten tips of (310), (111), and (100)-orientations. The topographic images generated from both the electron intensity variations and the field emission current indicate higher resolution capabilities with decreasing tip work function than with polycrystalline tungsten tips. The confinement of the electron beam transcends the resolution limitations of the geometrical models, which are determined by the minimum beam width.

Spin-polarised electrons in a one-magnet-only Mott spin junction.

The current flowing through a Mott spin junction depends on the relative spin orientation of the two ferromagnetic layers comprising the "source" and "drain" sides of the junction. The resulting current asymmetry is detected as giant or tunnelling magnetoresistance depending on whether the two ferromagnets are separated by a metal or an insulator. Based on the fundamental principles of reciprocity for spin-dependent electron scattering, one can envisage a one-magnet-only spin junction in which the source is non-magnetic, and the spin information is encoded by the spin polarisation of the electrons that have crossed or are backscattered from the drain magnetic layer. The practical significance of using an unpolarised source is that the state of the magnetic layer can be modified without affecting the process of probing it. Whether this reciprocity is realised in the actual junctions is not yet known. Here, we demonstrate a nano-sized, one-magnet-only Mott spin junction by measuring the finite spin polarisation of the backscattered electrons. Based on this finding, we conclude that since the junction acts as a spin filter, the magnetic layer must experience a spin transfer that could become detectable in view of the high current densities achievable in this technology.

Critical exponents and scaling invariance in the absence of a critical point.

The paramagnetic-to-ferromagnetic phase transition is classified as a critical phenomenon due to the power-law behaviour shown by thermodynamic observables when the Curie point is approached. Here we report the observation of such a behaviour over extraordinarily many decades of suitable scaling variables in ultrathin Fe films, for certain ranges of temperature T and applied field B. This despite the fact that the underlying critical point is practically unreachable because protected by a phase with a modulated domain structure, induced by the dipole-dipole interaction. The modulated structure has a well-defined spatial period and is realized in a portion of the (T, B) plane that extends above the putative critical temperature, where thermodynamic quantities do not display any singularity. Our results imply that scaling behaviour of macroscopic observables is compatible with an avoided critical point.

Thirty per cent contrast in secondary-electron imaging by scanning field-emission microscopy.

We perform scanning tunnelling microscopy (STM) in a regime where primary electrons are field-emitted from the tip and excite secondary electrons out of the target-the scanning field-emission microscopy regime (SFM). In the SFM mode, a secondary-electron contrast as high as 30% is observed when imaging a monoatomic step between a clean W(110)- and an Fe-covered W(110)-terrace. This is a figure of contrast comparable to STM. The apparent width of the monoatomic step attains the 1 nm mark, i.e. it is only marginally worse than the corresponding width observed in STM. The origin of the unexpected strong contrast in SFM is the material dependence of the secondary-electron yield and not the dependence of the transported current on the tip-target distance, typical of STM: accordingly, we expect that a technology combining STM and SFM will highlight complementary aspects of a surface while simultaneously making electrons, selected with nanometre spatial precision, available to a macroscopic environment for further processing.

Replication protein A induces the unwinding of long double-stranded DNA regions.

We have investigated nucleoprotein filaments composed of human replication protein A (RPA) and DNA by electron microscopy. At low ionic strengths, RPA complexes with single-stranded DNA are similar in length to protein-free DNA suggesting that RPA-bound DNA remains in an extended configuration under these conditions. However, severe compaction of RPA-DNA complexes occurs in buffers with > 2 mM MgCl2 or with 100 mM NaCl. At low ionic strengths, RPA binds to A + T-rich internal regions of linear double-stranded simian virus 40 (SV40) DNA and induces separation of complementary DNA strands. RPA also binds to closed-circular SV40 DNA, but requires the function of a DNA topoisomerase to invade and completely unwind duplex DNA regions. The ability of RPA to unwind long stretches of double-stranded DNA is not shared by the bacterial single-strand binding protein and the phage T4 gene 32 protein.

The role of the adsorption complex in the termination of filamentous phage assembly.

The adsorption complex of filamentous phage fd consists of two minor coat proteins, g3p and g6p, and is considered to be not only a structural entity, but also a functional unit to terminate phage assembly. Cells were infected with phage M13am8H1, which cannot assemble because it lacks the major coat protein g8p, although producing all of the other minor coat proteins. The membranes of infected cells were solubilized and analysed by non-denaturing PAGE and gel filtration. The data suggest the presence of the adsorption complex in these membranes. Furthermore, the non-polar gene 3 amber-mutant phage R171 was shown to lack g6p in the phage coat as well. The termination of assembly of this phage is disturbed, resulting in synthesis of polyphages. Electron micrographs and transient electrical birefringence show that these polyphages are eight times longer as compared to unit length phage. From these results, we conclude that the formation of the g3p-g6p complex is essential for correct termination of filamentous phage assembly.

Scaling theory of electric-field-assisted tunnelling.

Recent experiments report the current (I) versus voltage (V) characteristics of a tunnel junction consisting of a metallic tip placed at a distance d from a planar electrode, d varying over six orders of magnitude, from few nanometres to few millimetres. In the 'electric-field-assisted' (or 'field emission') regime, as opposed to the direct tunnelling regime used in conventional scanning tunnelling microscopy, all I-V curves are found to collapse onto one single graph when d is suitably rescaled, suggesting that the current I=I(V,d) is in reality a generalized homogeneous function of one single variable, i.e. [Formula: see text], where λ being some characteristic exponent and [Formula: see text] being a scaling function. In this paper, we provide a comprehensive explanation-based on analytical arguments, numerical simulations and further experimental results-for the scaling behaviour that we show to emerge for a variety of tip-plane geometries and thus seems to be a general feature of electric-field-assisted tunnelling.

Unwinding of chromatin by the SV40 large T antigen DNA helicase.

We have analysed the unwinding of nucleosomally organized DNA by simian virus 40 large tumour (T) antigen. Isolated T antigen can bind to existing nucleosome cores containing the viral replication origin sequence, which results in displacement of the histone octamer and unwinding of the DNA. However, specific binding to nucleosome cores is salt sensitive and nearly completely blocked under ionic conditions that otherwise support DNA replication. Once started, the progressing T antigen helicase, like an elongating RNA polymerase, is not further repressed by histone octamers, irrespective of the presence or absence of linker histone H1. Disruption of the nucleosomal structure in the process of unwinding may be assisted by the demonstrated interaction of the hexameric T antigen complex with histone proteins H1 and H3. Finally, our studies reveal the inability of topoisomerase I and/or II to continually relieve the superhelical tension of covalently closed circular minichromosomes as generated during their unwinding by T antigen. This may indicate that chromatin relaxation during the process of DNA replication can only be efficiently performed by a topoisomerase that is (trans)activated by other factors.

p53-catalyzed annealing of complementary single-stranded nucleic acids.

p53 has been reported to inhibit the DNA helicase intrinsic to simian virus 40 large tumor antigen (T antigen). We found that inhibition is not restricted to T antigen, but also affects several other DNA and RNA helicases. Complexing of the helicases by the p53 protein as a possible inactivation mechanism could be excluded. Instead, the anti-helicase activity can be explained by our finding that p53 binds with high affinity to single-stranded nucleic acids and has a strong DNA.DNA and RNA.RNA annealing activity. We could also show that p53 is able to alter the secondary structure of RNA and/or to influence dynamic RNA-RNA interactions. These results, and the fact that the affinity of p53 to RNA is about one order of magnitude higher than to single-stranded DNA, imply an RNA-specific function of p53 in vivo.

The interaction of SV40 large T antigen with unspecific double-stranded DNA: an electron microscopic study.

T antigen, an early protein encoded by simian virus 40 (SV40), is a specific DNA-binding protein with high affinity for elements in the viral origin of replication where it forms a double-hexameric complex as a prerequisite for DNA untwisting and, in the presence of ATP hydrolysis, for DNA unwinding. Like other specific DNA-binding proteins, T antigen also associates with DNA strands of random sequence albeit at reduced affinity. In addition, T antigen is able to unwind unspecific DNA sequences starting from internal binding sites. This property could be a step in the pathway leading to the chromosomal rearrangements that are frequently observed in SV40-transformed cells. This possibility prompted us to investigate the binding of T antigen to unspecific DNA using electron microscopy. We observed that the protein binds randomly to many unspecific DNA sites excluding a preference for particular DNA sequences or structural features. Addition of ATP to the binding buffer induces the formation of oligomeric, possibly hexameric, T antigen complexes that frequently align to form long arrays of DNA-bound protein. Magnesium salts induce the formation of tightly packed T antigen aggregates which bind to DNA to form many DNA branches and loops that emanate from the aggregated protein core. Upon ATP hydrolysis, aggregated T antigen catalyzes the unwinding of DNA duplices.

Genome structure of a virus infecting the marine brown alga Ectocarpus siliculosus.

We describe a procedure for the isolation of virus particles from the marine brown alga Ectocarpus siliculosus. Virus particles are composed of at least 13 different polypeptides, including two glycoproteins, and double-stranded DNA. A typical virus DNA preparation contains three fractions, namely linear DNA and circular DNA, each composed of about 320 kilobase pairs, as well as DNA fragments, 10 to 60 kilobase pairs in size. The large linear and the circular DNA contain single-stranded regions (average length: 2.9 kilobase). We propose that the native Ectocarpus virus genome is a circular DNA molecule whose double strand is interrupted by single-stranded regions. During the preparation procedure, the DNA circles tend to break at the single-stranded sites producing large linear as well as fragmented DNA.

Nucleic-acid-binding properties of hnRNP-U/SAF-A, a nuclear-matrix protein which binds DNA and RNA in vivo and in vitro.

We show that SAF-A, a nuclear protein which specifically binds vertebrate scaffold-attachment-region (SAR) elements with high affinity is identical with hnRNP-U, assumed to be involved in packaging of hnRNA in ribonucleoprotein particles. Ultraviolet cross-linking experiments show that the protein, referred to as hnRNP-U/SAF-A, is bound to chromosomal DNA in vivo. In vitro, the isolated protein binds to double-stranded and single-stranded DNA and forms higher ordered nucleic-acid-protein complexes. Filter-binding experiments performed with different types of natural and synthetic nucleic acids as substrates show that the protein binds DNA and RNA with different affinities and most likely at different binding sites. We conclude that hnRNP-U/SAF-A thus may have functions in the organisation of chromosomal DNA in addition to its suggested role in hnRNA metabolism.

Characterization of SAF-A, a novel nuclear DNA binding protein from HeLa cells with high affinity for nuclear matrix/scaffold attachment DNA elements.

We identified four proteins in nuclear extracts from HeLa cells which specifically bind to a scaffold attachment region (SAR) element from the human genome. Of these four proteins, SAF-A (scaffold attachment factor A), shows the highest affinity for several homologous and heterologous SAR elements from vertebrate cells. SAF-A is an abundant nuclear protein and a constituent of the nuclear matrix and scaffold. The homogeneously purified protein is a novel double stranded DNA binding protein with an apparent molecular weight of 120 kDa. SAF-A binds at multiple sites to the human SAR element; competition studies with synthetic polynucleotides indicate that these sites most probably reside in the multitude of A/T-stretches which are distributed throughout this element. In addition we show by electron microscopy that the protein forms large aggregates and mediates the formation of looped DNA structures.