Structural characterization of a new crystal form of the four-way Holliday junction formed by the DNA sequence d(CCGGTACCGG)2: sequence versus lattice?

DNA-strand exchange is a vital step in the recombination process, of which a key intermediate is the four-way DNA Holliday junction formed transiently in most living organisms. Here, the single-crystal structure at a resolution of 2.35 Å of such a DNA junction formed by d(CCGGTACCGG)2, which has crystallized in a more highly symmetrical packing mode to that previously observed for the same sequence, is presented. In this case, the structure is isomorphous to the mismatch sequence d(CCGGGACCGG)2, which reveals the roles of both lattice and DNA sequence in determining the junction geometry. The helices cross at the larger angle of 43.0° (the previously observed angle for this sequence was 41.4°) as a right-handed X. No metal cations were observed; the crystals were grown in the presence of only group I counter-cations.

Miniature transposable sequences are frequently mobilized in the bacterial plant pathogen Pseudomonas syringae pv. phaseolicola

Mobile genetic elements are widespread in Pseudomonas syringae, and often associate with virulence genes. Genome reannotation of the model bean pathogen P. syringae pv. phaseolicola 1448A identified seventeen types of insertion sequences and two miniature inverted-repeat transposable elements (MITEs) with a biased distribution, representing 2.8% of the chromosome, 25.8% of the 132-kb virulence plasmid and 2.7% of the 52-kb plasmid. Employing an entrapment vector containing sacB, we estimated that transposition frequency oscillated between 2.661025 and 1.161026, depending on the clone, although it was stable for each clone after consecutive transfers in culture media. Transposition frequency was similar for bacteria grown in rich or minimal media, and from cells recovered from compatible and incompatible plant hosts, indicating that growth conditions do not influence transposition in strain 1448A. Most of the entrapped insertions contained a full-length IS801 element, with the remaining insertions corresponding to sequences smaller than any transposable element identified in strain 1448A, and collectively identified as miniature sequences. From these, fragments of 229, 360 and 679-nt of the right end of IS801 ended in a consensus tetranucleotide and likely resulted from one-ended transposition of IS801. An average 0.7% of the insertions analyzed consisted of IS801 carrying a fragment of variable size from gene PSPPH_0008/PSPPH_0017, showing that IS801 can mobilize DNA in vivo. Retrospective analysis of complete plasmids and genomes of P. syringae suggests, however, that most fragments of IS801 are likely the result of reorganizations rather than one-ended transpositions, and that this element might preferentially contribute to genome flexibility by generating homologous regions of recombination. A further miniature sequence previously found to affect host range specificity and virulence, designated MITEPsy1 (100-nt), represented an average 2.4% of the total number of insertions entrapped in sacB, demonstrating for the first time the mobilization of a MITE in bacteria.

Fifty years of international business theory and beyond

As the field of international business has matured, there have been shifts in the core unit of analysis. First, there was analysis at country level, using national statistics on trade and foreign direct investment (FDI). Next, the focus shifted to the multinational enterprise (MNE) and the parent’s firm specific advantages (FSAs). Eventually the MNE was analysed as a network and the subsidiary became a unit of analysis. We untangle the last fifty years of international business theory using a classification by these three units of analysis. This is the country-specific advantage (CSA) and firm-specific advantage (FSA) matrix. Will this integrative framework continue to be useful in the future? We demonstrate that this is likely as the CSA/FSA matrix permits integration of potentially useful alternative units of analysis, including the broad region of the triad. Looking forward, we develop a new framework, visualized in two matrices, to show how distance really matters and how FSAs function in international business. Key to this are the concepts of compounded distance and resource recombination barriers facing MNEs when operating across national borders.

The role of double-stranded break repair in the creation of phenotypic diversity at cereal VRN1 loci

Nonhomologous repair of double-stranded breaks, although fundamental to the maintenance of genomic integrity in all eukaryotes, has received little attention as to its evolutionary consequences in the generation and selection of phenotypic diversity. Here we document the role of illegitimate recombination in the creation of novel alleles in VRN1 orthologs selected to confer adaptation to annual cropping systems in barley and wheat.

Construction of a Festuca pratensis BAC library for map-based cloning in Festulolium substitution lines

Introgression in Festulolium is a potentially powerful tool to isolate genes for a large number of traits which differ between Festuca pratensis Huds. and Lolium perenne L. Not only are hybrids between the two species fertile, but the two genomes can be distinguished by genomic in situ hybridisation and a high frequency of recombination occurs between homoeologous chromosomes and chromosome segments. By a programme of introgression and a series of backcrosses, L. perenne lines have been produced which contain small F. pratensis substitutions. This material is a rich source of polymorphic markers targeted towards any trait carried on the F. pratensis substitution not observed in the L. perenne background. We describe here the construction of an F. pratensis BAC library, which establishes the basis of a map-based cloning strategy in L. perenne. The library contains 49,152 clones, with an average insert size of 112 kbp, providing coverage of 2.5 haploid genome equivalents. We have screened the library for eight amplified fragment length polymorphism (AFLP) derived markers known to be linked to an F. pratensis gene introgressed into L. perenne and conferring a staygreen phenotype as a consequence of a mutation in primary chlorophyll catabolism. While for four of the markers it was possible to identify bacterial artificial chromosome (BAC) clones, the other four AFLPs were too repetitive to enable reliable identification of locus-specific BACs. Moreover, when the four BACs were partially sequenced, no obvious coding regions could be identified. This contrasted to BACs identified using cDNA sequences, when multiple genes were identified on the same BAC.

The linearly modulated IRSL red emission from feldspars

This paper uses the linear modulation technique to study red IRSL emission of potassium feldspars. Sub-samples were subjected to various pre-treatment and measurement conditions in an attempt to understand the relevant mechanisms of charge transfer. The linear modulation curves fitted most successfully to a sum of three first order components and we present supporting empirical evidence for the presence of three separate signal components. Additionally, the form of the red emission was observed to closely resemble the UV emission, implying the same donor charge concentrations may supply different recombination centres (assuming emission wavelength depends on centre type).

Chaos of Wolbachia Sequences Inside the Compact Fig Syconia of Ficus benjamina (Ficus: Moraceae)

Figs and fig wasps form a peculiar closed community in which the Ficus tree provides a compact syconium (inflorescence) habitat for the lives of a complex assemblage of Chalcidoid insects. These diverse fig wasp species have intimate ecological relationships within the closed world of the fig syconia. Previous surveys of Wolbachia, maternally inherited endosymbiotic bacteria that infect vast numbers of arthropod hosts, showed that fig wasps have some of the highest known incidences of Wolbachia amongst all insects. We ask whether the evolutionary patterns of Wolbachia sequences in this closed syconium community are different from those in the outside world. In the present study, we sampled all 17 fig wasp species living on Ficus benjamina, covering 4 families, 6 subfamilies, and 8 genera of wasps. We made a thorough survey of Wolbachia infection patterns and studied evolutionary patterns in wsp (Wolbachia Surface Protein) sequences. We find evidence for high infection incidences, frequent recombination between Wolbachia strains, and considerable horizontal transfer, suggesting rapid evolution of Wolbachia sequences within the syconium community. Though the fig wasps have relatively limited contact with outside world, Wolbachia may be introduced to the syconium community via horizontal transmission by fig wasps species that have winged males and visit the syconia earlier.

Direct probing of photoinduced electron transfer in a self- assembled biomimetic [2Fe2S]-hydrogenase complex using ultrafast vibrational spectroscopy

A pyridyl-functionalized diiron dithiolate complex, [μ-(4-pyCH2−NMI-S2)Fe2(CO)6] (3, py = pyridine(ligand), NMI = naphthalene monoimide) was synthesized and fully characterized. In the presence of zinc tetraphenylporphyrin (ZnTPP), a self-assembled 3·ZnTPP complex was readily formed in CH2Cl2 by the coordination of the pyridyl nitrogen to the porphyrin zinc center. Ultrafast photoinduced electron transfer from excited ZnTPP to complex 3 in the supramolecular assembly was observed in real time by monitoring the ν(CO) and ν(CO)NMI spectral changes with femtosecond time-resolved infrared (TRIR) spectroscopy. We have confirmed that photoinduced charge separation produced the monoreduced species by comparing the time-resolved IR spectra with the conventional IR spectra of 3•− generated by reversible electrochemical reduction. The lifetimes for the charge separation and charge recombination processes were found to be τCS = 40 ± 3 ps and τCR = 205 ± 14 ps, respectively. The charge recombination is much slower than that in an analogous covalent complex, demonstrating the potential of a supramolecular approach to extend the lifetime of the chargeseparated state in photocatalytic complexes. The observed vibrational frequency shifts provide a very sensitive probe of the delocalization of the electron-spin density over the different parts of the Fe2S2 complex. The TR and spectro-electrochemical IR spectra, electron paramagnetic resonance spectra, and density functional theory calculations all show that the spin density in 3•− is delocalized over the diiron core and the NMI bridge. This delocalization explains why the complex exhibits low catalytic dihydrogen production even though it features a very efficient photoinduced electron transfer. The ultrafast porphyrin-to-NMIS2−Fe2(CO)6 photoinduced electron transfer is the first reported example of a supramolecular Fe2S2-hydrogenase model studied by femtosecond TRIR spectroscopy. Our results show that TRIR spectroscopy is a powerful tool to investigate photoinduced electron transfer in potential dihydrogen-producing catalytic complexes, and that way to optimize their performance by rational approaches.

An eight-parent multiparent advanced generation inter-cross population for winter-sown wheat: creation, properties, and validation

MAGIC populations represent one of a new generation of crop genetic mapping resources combining high genetic recombination and diversity. We describe the creation and validation of an eight-parent MAGIC population consisting of 1091 F7 lines of winter-sown wheat (Triticum aestivum L.). Analyses based on genotypes from a 90,000-single nucleotide polymorphism (SNP) array find the population to be well-suited as a platform for fine-mapping quantitative trait loci (QTL) and gene isolation. Patterns of linkage disequilibrium (LD) show the population to be highly recombined; genetic marker diversity among the founders was 74% of that captured in a larger set of 64 wheat varieties, and 54% of SNPs segregating among the 64 lines also segregated among the eight founder lines. In contrast, a commonly used reference bi-parental population had only 54% of the diversity of the 64 varieties with 27% of SNPs segregating. We demonstrate the potential of this MAGIC resource by identifying a highly diagnostic marker for the morphological character "awn presence/absence" and independently validate it in an association-mapping panel. These analyses show this large, diverse, and highly recombined MAGIC population to be a powerful resource for the genetic dissection of target traits in wheat, and it is well-placed to efficiently exploit ongoing advances in phenomics and genomics. Genetic marker and trait data, together with instructions for access to seed, are available at http://www.niab.com/MAGIC/.

Nicked-site substrates for a serine recombinase reveal enzyme–DNA communications and an essential tethering role of covalent enzyme–DNA linkages

To analyse the mechanism and kinetics of DNA strand cleavages catalysed by the serine recombinase Tn3 resolvase, we made modified recombination sites with a single-strand nick in one of the two DNA strands. Resolvase acting on these sites cleaves the intact strand very rapidly, giving an abnormal half-site product which accumulates. We propose that these reactions mimic second-strand cleavage of an unmodified site. Cleavage occurs in a synapse of two sites, held together by a resolvase tetramer; cleavage at one site stimulates cleavage at the partner site. After cleavage of a nicked-site substrate, the half-site that is not covalently linked to a resolvase subunit dissociates rapidly from the synapse, destabilizing the entire complex. The covalent resolvase–DNA linkages in the natural reaction intermediate thus perform an essential DNA-tethering function. Chemical modifications of a nicked-site substrate at the positions of the scissile phosphodiesters result in abolition or inhibition of resolvase-mediated cleavage and effects on resolvase binding and synapsis, providing insight into the serine recombinase catalytic mechanism and how resolvase interacts with the substrate DNA.

Fungicide-resistance management tactics: impacts on Zymoseptoria tritici populations

Azoles and Succinate Dehydrogenase Inhibitors (SDHIs) are the main fungicides available for septoria tritici blotch control, causal agent Zymoseptoria tritici. Decline in azole sensitivity, in combination with European legislation, poses a threat to wheat production in Ireland. Azole fungicides select CYP51 mutations differentially; it was hypothesised that using combinations of azoles could be an effective anti-resistance tool. Naturally inoculated field experiments were carried out in order to understand the impacts of using combinations of azoles, epoxiconazole and metconazole, on azole sensitivity. Approximately 3700 isolates were isolated and their sensitivity to both azoles analysed. Findings showed that limiting the number of applications, by alternating each fungicide, slowed selection for reduced azole sensitivity. Limiting azole use by reducing doses did not reduce selection for decreased azole sensitivity. Although not complete, cross-resistance was observed between the two azoles, which will lead to general reduction in azole sensitivity. A sub-selection of isolates from each treatment at each location were analysed for changes in the CYP51 gene. Sequence analysis identified 49 combinations of mutations in the CYP51 gene, and three different inserts in the CYP51 promoter. Intragenic recombination also featured in these populations. Baseline studies of five new SDHIs were carried out on 209 naturally infected, non-SDHI-treated isolates. With the exception of fluopyram, cross-resistance was apparent between the SDHIs. Analysis of 2300 isolates found that when compared to the solo products, mixing the SDHI isopyrazam and the azole epoxiconazole increased epoxiconazole sensitivity, but had no apparent effect on isopyrazam sensitivity. SDHI resistance-conferring mutations were absent in the baseline and experimental isolates. As long as azoles are used, Z. tritici populations will continue to evolve towards resistance. Combining different modes-of-action, SDHIs and multi-sites, with azoles will relieve some of that selective pressure. To get the best out of available fungicides, they should be used in combination with host resistance and good crop management practices.

Rethinking the literature on the performance of Chinese multinational enterprises

We synthesize the literature on Chinese multinational enterprises (MNEs) and find that much of the prior research is based on as few as a dozen case studies of Chinese firms. They are so case-specific that it has led to a misplaced call for new theories to explain Chinese firms’ internationalization. In an attempt to better relate theory with empirical evidence, we examine the largest 500 Chinese manufacturing firms. We aim to find out the number of Chinese manufacturing firms to be true MNEs by definition, and to examine their financial performance relative to global peers using the financial benchmarking method. We develop our theoretical perspectives from new internalization theory. We find that there are only 49 Chinese manufacturing firms to be true MNEs, whereas the rest is purely domestic firms. Their performance is poor relative to global peers. Chinese MNEs have home country bound firm-specific advantages (FSAs), which are built upon home country-specific advantages (home CSAs). They have not yet developed advanced management capabilities through recombination with host CSAs. Essentially, they acquire foreign firms to increase their sales in domestic market, but they fail to be competitive internationally and to achieve superior performance in overseas operations. Our findings have important strategic implications for managers, public policy makers, and academic research.