Correcting errors in synthetic DNA through consensus shuffling

Although efficient methods exist to assemble synthetic oligonucleotides into genes and genomes, these suffer from the presence of 1–3 random errors/kb of DNA. Here, we introduce a new method termed consensus shuffling and demonstrate its use to significantly reduce random errors in synthetic DNA. In this method, errors are revealed as mismatches by re-hybridization of the population. The DNA is fragmented, and mismatched fragments are removed upon binding to an immobilized mismatch binding protein (MutS). PCR assembly of the remaining fragments yields a new population of full-length sequences enriched for the consensus sequence of the input population. We show that two iterations of consensus shuffling improved a population of synthetic green fluorescent protein (GFPuv) clones from ∼60 to >90% fluorescent, and decreased errors 3.5- to 4.3-fold to final values of ∼1 error per 3500 bp. In addition, two iterations of consensus shuffling corrected a population of GFPuv clones where all members were non-functional, to a population where 82% of clones were fluorescent. Consensus shuffling should facilitate the rapid and accurate synthesis of long DNA sequences

Computation of protein geometry and its applications: Packing and function prediction

This chapter discusses geometric models of biomolecules and geometric constructs, including the union of ball model, the weigthed Voronoi diagram, the weighted Delaunay triangulation, and the alpha shapes. These geometric constructs enable fast and analytical computaton of shapes of biomoleculres (including features such as voids and pockets) and metric properties (such as area and volume). The algorithms of Delaunay triangulation, computation of voids and pockets, as well volume/area computation are also described. In addition, applications in packing analysis of protein structures and protein function prediction are also discussed.Comment: 32 pages, 9 figure

Tropospheric distribution of sulphate aerosol mass and number concentration during INDOEX-IFP and its transport over the Indian Ocean: a GCM study

International audienceAn interactive sulphate aerosol chemistry module has been incorporated in the Laboratoire de Météorologie Dynamique General Circulation Model (LMD-GCM) to simulate the sulphur chemistry during the Indian Ocean Experiment (INDOEX) Intensive Field Phase-1999 (INDOEX-IFP). The originality of this module is its ability to predict particle mass and number concentration for the Aitken and accumulation modes. The model qualitatively reproduces the spatial patterns of observations on sulphate aerosol during INDOEX. On the basis of size distribution retrieved from the observations made along the cruise route during 1998 and 1999, the model successfully simulates the order of magnitude and the general north-south gradient in aerosol number concentration. The result shows the southward migration of minimum concentrations, which follows ITCZ (Inter Tropical Convergence Zone) migration. Sulphate surface concentration during INDOEX-IFP at Kaashidhoo (73.46° E, 4.96° N) gives an agreement within a factor of 2 to 3. Predicted sulphate aerosol optical depth (AOD) matches reasonably with measured values, indicating the capability of this model to predict the vertically integrated column sulphate burden. The Indian contribution to estimated sulphate burden over India is more than 60% with values upto 40% over the Arabian Sea

WRF-CMAQ two-way coupled system with aerosol feedback: software development and preliminary results

Air quality models such as the EPA Community Multiscale Air Quality (CMAQ) require meteorological data as part of the input to drive the chemistry and transport simulation. The Meteorology-Chemistry Interface Processor (MCIP) is used to convert meteorological data into CMAQ-ready input. Key shortcoming of such one-way coupling include: excessive temporal interpolation of coarsely saved meteorological input and lack of feedback of atmospheric pollutant loading on simulated dynamics. We have developed a two-way coupled system to address these issues. A single source code principle was used to construct this two-way coupling system so that CMAQ can be consistently executed as a stand-alone model or part of the coupled system without any code changes; this approach eliminates maintenance of separate code versions for the coupled and uncoupled systems. The design also provides the flexibility to permit users: (1) to adjust the call frequency of WRF and CMAQ to balance the accuracy of the simulation versus computational intensity of the system, and (2) to execute the two-way coupling system with feedbacks to study the effect of gases and aerosols on short wave radiation and subsequent simulated dynamics. Details on the development and implementation of this two-way coupled system are provided. When the coupled system is executed without radiative feedback, computational time is virtually identical when using the Community Atmospheric Model (CAM) radiation option and a slightly increased (~8.5 %) when using the Rapid Radiative Transfer Model for GCMs (RRTMG) radiation option in the coupled system compared to the offline WRF-CMAQ system. Once the feedback mechanism is turned on, the execution time increases only slightly with CAM but increases about 60 % with RRTMG due to the use of a more detailed Mie calculation in this implementation of feedback mechanism. This two-way model with radiative feedback shows noticeably reduced bias in simulated surface shortwave radiation and 2 m temperatures as well improved correlation of simulated ambient ozone and PM<sub>2.5</sub> relative to observed values for a test case with significant tropospheric aerosol loading from California wildfires

A plume-in-grid approach to characterize air quality impacts of aircraft emissions at the Hartsfield-Jackson Atlanta International Airport

This study examined the impacts of aircraft emissions during the landing and takeoff cycle on PM2.5 concentrations during the months of June 2002 and July 2002 at the Hartsfield-Jackson Atlanta International Airport. Primary and secondary pollutants were modeled using the Advanced Modeling System for Transport, Emissions, Reactions, and Deposition of Atmospheric Matter (AMSTERDAM). AMSTERDAM is a modified version of the Community Multiscale Air Quality (CMAQ) model that incorporates a plume-in-grid process to simulate emissions sources of interest at a finer scale than can be achieved using CMAQ's model grid. Three fundamental issues were investigated: the effects of aircraft on PM2.5 concentrations throughout northern Georgia, the differences resulting from use of AMSTERDAM's plume-in-grid process rather than a traditional CMAQ simulation, and the concentrations observed in aircraft plumes at sub-grid scales. Comparison of model results with an air quality monitor located in the vicinity of the airport found that normalized mean bias ranges from -77.5% to 6.2% and normalized mean error ranges from 40.4% to 77.5%, varying by species. Aircraft influence average PM2.5 concentrations by up to 0.232 μg m-3 near the airport and by 0.001-0.007 μg m-3 throughout the Atlanta metro area. The plume-in-grid process increases concentrations of secondary PM pollutants by 0.005-0.020 μg m-3 (compared to the traditional grid-based treatment) but reduces the concentration of non-reactive primary PM pollutants by up to 0.010 μg m-3, with changes concentrated near the airport. Examination of sub-grid scale results indicates that puffs within 20 km of the airport often have average PM2.5 concentrations one order of magnitude higher than aircraft contribution to the grid cells containing those puffs, and within 1-4 km of emitters, puffs may have PM2.5 concentrations 3 orders of magnitude greater than the aircraft contribution to their grid cells. 21% of all aircraft-related puffs from the Atlanta airport have at least 0.1 μg m-3 PM2.5 concentrations. Median daily puff concentrations vary between 0.017 and 0.134 μg m-3, while maximum daily puff concentrations vary between 6.1 and 42.1 μg m-3 during the 2-month period. In contrast, median daily grid concentrations vary between 0.015 and 0.091 μg m-3, while maximum daily grid concentrations vary between 0.751 and 2.55 μg m-3. Future researchers may consider using AMSTERDAM to understand the impacts of aircraft emissions at other airports, for proposed future airports, for airport expansion projects under various future scenarios, and for other national-scale studies specifically when the maximum impacts at fine scales are of interest

Modeling the impact of sea-spray on particle concentrations in a coastal city

Abstract 18 An atmospheric chemistry-transport model is used to assess the impacts of sea-spray chemistry 19 on the particle composition in and downwind of a coastal city -Vancouver, British Columbia. 20 Reactions in/on sea-spray affect the entire particle ensemble and particularly the size distribution 21 of particle nitrate. 2

Modeling the impact of sea-spray on particle concentrations in a coastal city

An atmospheric chemistry-transport model is used to assess the impacts of sea-spray chemistry on the particle composition in and downwind of a coastal city--Vancouver, British Columbia. Reactions in/on sea-spray affect the entire particle ensemble and particularly the size distribution of particle nitrate. Urban air quality, and particularly airborne particles, is a major concern in terms of human health impacts. Sea-spray is known to be a major component of the particle ensemble at coastal sites yet relatively few air quality models include the interaction of gases with sea-spray and the fate of the particles produced. Sea-spray is not an inert addition to the particle ensemble because heterogeneous chemistry in/on sea-spray droplets changes the droplets composition and the particle size distribution, which impacts deposition and the ion balance in different particle size fractions. It is shown that the ISOPART model is capable of simulating gas and particle concentrations in the coastal metropolis of Vancouver and the surrounding valley. It is also demonstrated that to accurately simulate ambient concentrations of particles and reactive/soluble gases in a coastal valley it is absolutely critical to include heterogeneous chemistry in/on sea-spray. Partitioning of total particle-NO{sub 3}{sup -} between sea-spray and NH{sub 4}NO{sub 3} is highly sensitive to the amount of sea-spray present, and hence the initial vertical profile, sea-spray source functions [48] and the wind speed. When a fixed wind speed is used to initialize the sea-spray vertical profiles, as expected, the sea-spray concentration decays with distance inland, but the particle-NO{sub 3}{sup -} concentration decays more slowly because it is also a function of the uptake rate for HNO{sub 3}. The simulation results imply model analyses of air quality in coastal cities conducted without inclusion of sea-spray interactions may yield highly misleading results in terms of emission sensitivities of the PM size distribution. The sensitivity of the model results to the initial sea spray profile further suggests there would be great benefit in better definition of the vertical profile of size resolved sea-spray for use in such model studies

High throughput mutagenesis for identification of residues regulating human prostacyclin (hIP) receptor

The human prostacyclin receptor (hIP receptor) is a seven-transmembrane G protein-coupled receptor (GPCR) that plays a critical role in vascular smooth muscle relaxation and platelet aggregation. hIP receptor dysfunction has been implicated in numerous cardiovascular abnormalities, including myocardial infarction, hypertension, thrombosis and atherosclerosis. Genomic sequencing has discovered several genetic variations in the PTGIR gene coding for hIP receptor, however, its structure-function relationship has not been sufficiently explored. Here we set out to investigate the applicability of high throughput random mutagenesis to study the structure-function relationship of hIP receptor. While chemical mutagenesis was not suitable to generate a mutagenesis library with sufficient coverage, our data demonstrate error-prone PCR (epPCR) mediated mutagenesis as a valuable method for the unbiased screening of residues regulating hIP receptor function and expression. Here we describe the generation and functional characterization of an epPCR derived mutagenesis library compromising >4000 mutants of the hIP receptor. We introduce next generation sequencing as a useful tool to validate the quality of mutagenesis libraries by providing information about the coverage, mutation rate and mutational bias. We identified 18 mutants of the hIP receptor that were expressed at the cell surface, but demonstrated impaired receptor function. A total of 38 non-synonymous mutations were identified within the coding region of the hIP receptor, mapping to 36 distinct residues, including several mutations previously reported to affect the signaling of the hIP receptor. Thus, our data demonstrates epPCR mediated random mutagenesis as a valuable and practical method to study the structurefunction relationship of GPCRs. © 2014 Bill et al

WRF-CMAQ two-way coupled system with aerosol feedback: software development and preliminary results

Air quality models such as the EPA Community Multiscale Air Quality (CMAQ) require meteorological data as part of the input to drive the chemistry and transport simulation. The Meteorology-Chemistry Interface Processor (MCIP) is used to convert meteorological data into CMAQ-ready input. Key shortcoming of such one-way coupling include: excessive temporal interpolation of coarsely saved meteorological input and lack of feedback of atmospheric pollutant loading on simulated dynamics. We have developed a two-way coupled system to address these issues. A single source code principle was used to construct this two-way coupling system so that CMAQ can be consistently executed as a stand-alone model or part of the coupled system without any code changes; this approach eliminates maintenance of separate code versions for the coupled and uncoupled systems. The design also provides the flexibility to permit users: (1) to adjust the call frequency of WRF and CMAQ to balance the accuracy of the simulation versus computational intensity of the system, and (2) to execute the two-way coupling system with feedbacks to study the effect of gases and aerosols on short wave radiation and subsequent simulated dynamics. Details on the development and implementation of this two-way coupled system are provided. When the coupled system is executed without radiative feedback, computational time is virtually identical when using the Community Atmospheric Model (CAM) radiation option and a slightly increased (~8.5%) when using the Rapid Radiative Transfer Model for GCMs (RRTMG) radiation option in the coupled system compared to the offline WRF-CMAQ system. Once the feedback mechanism is turned on, the execution time increases only slightly with CAM but increases about 60% with RRTMG due to the use of a more detailed Mie calculation in this implementation of feedback mechanism. This two-way model with radiative feedback shows noticeably reduced bias in simulated surface shortwave radiation and 2-m temperatures as well improved correlation of simulated ambient ozone and PM2.5 relative to observed values for a test case with significant tropospheric aerosol loading from California wildfires

A 41,500 year-old decorated ivory pendant from Stajnia Cave (Poland)

Evidence of mobiliary art and body augmentation are associated with the cultural innovations introduced by Homo sapiens at the beginning of the Upper Paleolithic. Here, we report the discovery of the oldest known human-modified punctate ornament, a decorated ivory pendant from the Paleolithic layers at Stajnia Cave in Poland. We describe the features of this unique piece, as well as the stratigraphic context and the details of its chronometric dating. The Stajnia Cave plate is a personal 'jewellery' object that was created 41,500 calendar years ago (directly radiocarbon dated). It is the oldest known of its kind in Eurasia and it establishes a new starting date for a tradition directly connected to the spread of modern Homo sapiens in Europe