Rapid metabolic pathway assembly and modification using serine integrase site-specific recombination

Synthetic biology requires effective methods to assemble DNA parts into devices and to modify these devices once made. Here we demonstrate a convenient rapid procedure for DNA fragment assembly using site-specific recombination by ϕC31 integrase. Using six orthogonal attP/attB recombination site pairs with different overlap sequences, we can assemble up to five DNA fragments in a defined order and insert them into a plasmid vector in a single recombination reaction. ϕC31 integrase-mediated assembly is highly efficient, allowing production of large libraries suitable for combinatorial gene assembly strategies. The resultant assemblies contain arrays of DNA cassettes separated by recombination sites, which can be used to manipulate the assembly by further recombination. We illustrate the utility of these procedures to (i) assemble functional metabolic pathways containing three, four or five genes; (ii) optimize productivity of two model metabolic pathways by combinatorial assembly with randomization of gene order or ribosome binding site strength; and (iii) modify an assembled metabolic pathway by gene replacement or addition

Safety evaluation of substituted thiophenes used as flavoring ingredients

AbstractThis publication is the second in a series by the Expert Panel of the Flavor and Extract Manufacturers Association summarizing the conclusions of its third systematic re-evaluation of the safety of flavorings previously considered to be generally recognized as safe (GRAS) under conditions of intended use. Re-evaluation of GRAS status for flavorings is based on updated considerations of exposure, structural analogy, metabolism, pharmacokinetics and toxicology and includes a comprehensive review of the scientific information on the flavorings and structurally related substances. Of the 12 substituted thiophenes reviewed here, 11 were reaffirmed as GRAS based on their rapid absorption, metabolism and excretion in humans and animals; the low estimated dietary exposure from flavor use; the wide margins of safety between the conservative estimates of intake and the no-observed-adverse effect levels; and the lack of significant genotoxic and mutagenic potential. For one of the substituted thiophenes, 3-acetyl-2,5-dimethylthiophene, it was concluded that more detailed exposure information, comparative metabolism studies and comprehensive toxicity data, including an in-depth evaluation of the mechanism of action for any adverse effects observed, are required for continuation of its FEMA GRAS™ status. In the absence of these data, the compound was removed from the FEMA GRAS list

Is null-point reconnection important for solar flux emergence?

The role of null-point reconnection in a 3D numerical MHD model of solar emerging flux is investigated. The model consists of a twisted magnetic flux tube rising through a stratified convection zone and atmosphere to interact and reconnect with a horizontal overlying magnetic field in the atmosphere. Null points appear as the reconnection begins and persist throughout the rest of the emergence, where they can be found mostly in the model photosphere and transition region, forming two loose clusters on either side of the emerging flux tube. Up to 26 nulls are present at any one time, and tracking in time shows that there is a total of 305 overall, despite the initial simplicity of the magnetic field configuration. We find evidence for the reality of the nulls in terms of their methods of creation and destruction, their balance of signs, their long lifetimes, and their geometrical stability. We then show that due to the low parallel electric fields associated with the nulls, null-point reconnection is not the main type of magnetic reconnection involved in the interaction of the newly emerged flux with the overlying field. However, the large number of nulls implies that the topological structure of the magnetic field must be very complex and the importance of reconnection along separators or separatrix surfaces for flux emergence cannot be ruled out.Comment: 26 pages, 12 figures. Added one referenc

Measurements of the t(t)Overbar charge asymmetry using the dilepton decay channel in pp collisions at root s=7 TeV

The tt¯ charge asymmetry in proton-proton collisions at s√ = 7 TeV is measured using the dilepton decay channel (ee, e μ , or μμ ). The data correspond to a total integrated luminosity of 5.0 fb −1 , collected by the CMS experiment at the LHC. The tt and lepton charge asymmetries, defined as the differences in absolute values of the rapidities between the reconstructed top quarks and antiquarks and of the pseudorapidities between the positive and negative leptons, respectively, are measured to be A C = −0 . 010 ± 0 . 017 (stat . ) ± 0 . 008 (syst . ) and AlepC = 0 . 009 ± 0 . 010 (stat . ) ± 0 . 006 (syst . ). The lepton charge asymmetry is also measured as a function of the invariant mass, rapidity, and transverse momentum of the tt¯ system. All measurements are consistent with the expectations of the standard model