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The Molecular Tagging (MT) technique is a promising methodology for locally measuring velocity and temperature fields in rarefied gas flows. Recently, Molecular Tagging Velocimetry (MTV) has been successfully applied to gas flows in mini-channels in the continuum regime at high pressure and early slip-flow regime at lower pressure. As the operating pressure decreases, diffusion effects become more pronounced, and in MTV, they hinder the extraction of the correct velocity profile by simply dividing the displacement profile of the tagged molecular line by time of flight. To address this issue, a reconstruction method that considers Taylor dispersion was previously developed to extract the velocity profile, considering the diffusion effects of the tracer molecules within the carrier gas. This reconstruction method successfully extracted the correct velocity profile in the continuum flow regime. However, the method still faces challenges in the slip-flow regime. Since there is currently no consensus in the literature regarding the kinetic diameter value of acetone vapor, the diffusion coefficient estimation is uncertain especially at low pressures. This is why, in this study, we propose an original optical method to measure the diffusion coefficient of acetone vapor. This is achieved by linking the temporal evolution of the spatial photoluminescence distribution of acetone vapor to the diffusion coefficient via the Chapman-Enskog theory. Our research provides measurements of these parameters for a wide range of pressures (0.5–10 kPa) at ambient temperature. [ABSTRACT FROM AUTHOR]

Long-lasting emission from femtosecond excitation of nitrogen-based flows shows promise as a useful mechanism for a molecular tagging velocimetry instrument. The technique, known as femtosecond laser electronic excitation tagging (FLEET), was invented at Princeton a decade ago and has quickly been adopted and used in a variety of high-speed ground test flow facilities. The short temporal scales offered by femtosecond amplifiers permit nonresonant multiphoton excitation, dissociation, and weak ionization of a gaseous medium near the beam's focus without the generation of a laser spark observed with nanosecond systems. Gated, intensified imaging of the resulting emission enables the tracking of tagged molecules, thereby measuring one to three components of velocity. Effects of local heating and acoustic disturbances can be mitigated with the selection of a shorter-wavelength excitation source. This review surveys the development of FLEET over the decade since its inception, as it has been implemented in several test facilities to make accurate, precise, and seedless velocimetry measurements for studying complex high-speed flows. [ABSTRACT FROM AUTHOR]

Aims: To conduct molecular tagging of the biocontrol fungus Trichoderma asperellum strain T4 and elucidate its colonization patterns in soil. Methods and Results: We constructed an expression vector harbouring a hygromycin B‐resistant gene (hph) and an efficient green fluorescent protein (egfp) gene. By applying Agrobacterium AGL‐1‐mediated genetic transformation technology, we conducted molecular tagging of T. asperellum and monitored the colonization dynamics of T. asperellum in soil. The results of tracking five independent transformants of T. asperellum indicated that its expansion rates ranged from 4·7 to 6·8 cm week−1. After inoculation in soil, the quantities of T. asperellum could be maintained at over 10 × 104 CFU per gram soil in the first year. In the third year after inoculation, the quantities of T. asperellum in soil were still higher than 1 × 103 CFU per gram soil. In addition, molecularly tagged T. asperellum in soil in the second year (i.e. 12 months) after inoculation could still reach the biocontrol effect on cucumber Rhizoctonia rot by more than 74%. Conclusion: Trichoderma asperellum strain T4 is capable of effectively colonizing in soil and surviving for more than 1 year. Significance and Impact of the Study: This study has provided the scientific basis for applying T. asperellum as the biocontrol fungus for prevention and control of plant diseases. [ABSTRACT FROM AUTHOR]

Helium-4 in the superfluid phase (He II) is a two-fluid system that exhibits fascinating quantum hydrodynamics with important scientific and engineering applications. However, the lack of high-precision flow measurement tools in He II has impeded the progress in understanding and utilizing its hydrodynamics. In recent years, there have been extensive efforts in developing quantitative flow visualization techniques applicable to He II. In particular, a powerful molecular tagging velocimetry (MTV) technique, based on tracking thin lines of He 2 ∗ excimer molecules created via femtosecond laser-field ionization in helium, has been developed in our laboratory. This technique allows unambiguous measurement of the normal fluid velocity field in the two-fluid system. Nevertheless, there are two limitations to this technique: (1) only the velocity component perpendicular to the tracer line can be measured; and (2) there is an inherent error in determining the perpendicular velocity. In this paper, we discuss how these issues can be resolved by advancing the MTV technique. We also discuss two novel schemes for tagging and producing He 2 ∗ tracers. The first method allows the creation of a tagged He 2 ∗ tracer line without the use of an expensive femtosecond laser. The second method enables full-space velocity field measurement through tracking small clouds of He 2 ∗ molecules created via neutron- 3 He absorption reactions in He II. [ABSTRACT FROM AUTHOR]

Pathogenic mutations in relapse-associated genes in pediatric acute lymphoblastic leukemia may improve risk stratification when detected at subclonal levels at primary diagnosis. However, to detect subclonal mutations upfront, a deep-sequencing approach with high specificity and sensitivity is required. Here, we performed a proof-of-principle study to detect low-level mosaic RAS pathway mutations by deep sequencing using random tagging-based single molecule Molecular Inversion Probes (smMIPs). The smMIP-based approach could sensitively detect variants with allele frequency as low as 0.4%, which could all be confirmed by other techniques. In comparison, with standard deep-sequencing techniques we reached a detection threshold of only 2.5%, which hampered detection of seven low-level mosaic mutations representing 24% of all detected mutations. We conclude that smMIP-based deep-sequencing outperforms standard deep-sequencing techniques by showing lower background noise and high specificity, and is the preferred technology for detecting mutations upfront, particularly in genes in which mutations show limited clustering in hotspots. [ABSTRACT FROM AUTHOR]

A molecular tagging technique utilizing evanescent wave illumination was developed to investigate the motion of a caged fluorescent dye in the vicinity of the microchannel wall surface in electroosmotic and pressure-driven flows. A line pattern in a buffer solution was written by a pulsed UV laser and the uncaged dye was excited by the evanescent wave with total internal reflection inside the glass wall using an objective lens. The velocities calculated by the measured displacement of the near-wall tagged region were compared with the results of molecular tagging using volume illumination, which represents the bulk flow information. Concerning electroosmotic flow, the micro-PIV technique using a confocal microscope system was applied to the microchannel rinsed by the caged fluorescein beforehand in comparison with a pure glass-PDMS microchannel to examine the effect of dye adsorption to the wall on the electroosmotic mobility. The electroosmotic mobility obtained by evanescent wave molecular tagging (EWMT) showed close to the micro-PIV measurement result near the glass wall for the rinsed case and the uncaged dye at the almost constant velocity remained in the depthwise illumination region. On the other hand, the dye velocity in pressure-driven flow by EWMT increased rapidly with respect to time. The uncaged dye convected to the streamwise direction dispersed toward the wall due to the concentration gradient of the dye, which was confirmed by the numerical simulations.

Abstract: Non-invasive genetic techniques have proven to be cost-effective for monitoring and studying otter populations, largely due to the linearity of otter territories and the marking behavior of the species. After a severe decline, in the 60, the Eurasian otter is recovering in the Iberian Peninsula. However, the recovery pattern is not homogeneous and the species is still considered “Threatened” in many regions. During 2007–2010 a systematic non-invasive genetic sampling effort was carried out to determine the spatial distribution and to estimate the population size of an endangered otter population in northern Iberian Peninsula (Basque Country). Samples were identified to species level by sequencing a 226bp mtDNA fragment prior to genotyping. Among the 132 obtained samples, 127 (98.4%) belonged to the study species, one sample was genetically identified as European mink (Mustela lutreola) and one as American mink (Neovison vison) while genetic species confirmation was not possible in the three remaining samples. These results provided novel and accurate data on species distribution, highlighting an overall increase of 25% in 10×10 UTM grids occupied by otter and a clear pattern of re-colonization upstream of the main rivers. All samples corresponding to otter were subsequently individually genotyped using a novel multiplex panel of 11 microsatellite markers and sexed by typing the sex-chromosome-related gene ZFX/ZFY. We obtained a complete individual genetic profile for 55 samples (genotyping success 43%), corresponding to 20 different individuals (11 females, 6 males, and 3 individuals of unknown gender). The mean otter density in occupied areas estimated to be 0.09 (0.06–0.12) individuals per river kilometer. The present study enabled us to obtain updated and relevant information about this elusive species’ distribution and population size, essential to define population status and to design successful and effective management and conservation programs. [Copyright &y& Elsevier]

Reliable estimates of population parameters are necessary for effective management and conservation actions. The use of genetic data for capture-recapture (CR) analyses has become an important tool to estimate population parameters for elusive species. Strong emphasis has been placed on the genetic analysis of non-invasive samples, or on the CR analysis; however, little attention has been paid to the simultaneous overview of the full non-invasive genetic CR analysis, and the important insights gained by understanding the interactions between the different parts of the technique. Here, we review the three main steps of the approach: designing the appropriate sampling scheme, conducting the genetic lab analysis, and applying the CR analysis to the genetic results; and present a synthesis of this topic with the aim of discussing the primary limitations and sources of error. We discuss the importance of the integration between these steps, the unique situations which occur with non-invasive studies, the role of ecologists and geneticists throughout the process, the problem of error propagation, and the sources of biases which can be present in the final estimates. We highlight the importance of team collaboration and offer a series of recommendations to wildlife ecologists who are not familiar with this topic yet but may want to use this tool to monitor populations through time. [ABSTRACT FROM AUTHOR]

• Composition of size-fractionated river NOM was characterized by Orbitrap-MS. • The correlations between HBQs and source water NOM components were established. • Molecular tagging revealed the contribution of functional groups on HBQs formation. • HBQs yield was strongly impacted by NOM molecular composition and functional groups. • 50 kDa source water NOM was more conducive to form HBQs during chlorination. Halobenzoquinones (HBQs) are emerging disinfection byproducts generated during the reaction of chlorine disinfectant with natural organic matter (NOM) in source water. In this study, the correlations between molecular weight and HBQs generation of river NOM was evaluated. The compositional and functional characteristics of primary HBQs precursors were revealed by using Orbitrap mass spectrometry combined with molecular tagging. The NOM fraction larger than 50 kDa resulted in approximately 9 times more HBQs (50.9 ± 2.7 ng/mgC) than low molecular weight fractions. Significant correlations were found between the yields of HBQs and lignin-like and highly oxygen compounds in NOM, suggesting their critical roles in HBQs formation. Derivatizating the aldehydes, ketones, hydroxyl and carboxyl groups in NOM could reduce HBQs yields by 90.7%–100%. Unraveling the molecular characteristics of HBQs precursors in NOM would greatly benefit the prediction of HBQs yields of different source water, and develop more efficient disinfection byproduct control strategies. [Display omitted] [ABSTRACT FROM AUTHOR]

The improvement of cotton fiber quality is extremely important because of changes in spinning technology. The identification of the stable QTLs affecting fiber traits across different generations will be greatly helpful to be used effectively in molecular marker-assisted selection to improve fiber quality of cotton cultivars in the future. Using three elite fiber lines of Upland cotton (Gossypium hirsutumL.) as parents, three linkage maps were constructed to tag QTLs for fiber qualities using SSR markers. There were 39 QTLs, 17 significant QTLs, LOD = 3.0 and 22 suggestive QTLs, 3.0>LOD = 2.0, detected by composite interval mapping for fiber traits, in which 11 QTLs were for fiber length, 10 for fiber strength, 9 for micronaire and 9 for fiber elongation. Out of 17 significant QTLs, 5 QTLs with high logarithm of odds (LOD) score value and stable effect could be found in both F2 and F2:3 segregating populations, showing a great potential for molecular-assisted selection in improving fiber quality. At least three common QTLs could be identified in two populations. These common QTLs detected in different populations suggested that there existed elite fiber genes and possibly of the same origin. In addition, we found three pairs of putative homoeologous QTLs,qFL-7-1candqFL-16-1c,qFS-D03-1a,qFS-A02-1bandqFS-A02-1c, andqFE-D03-1aandqFE-A02-1c. Our results provided a better understanding of the genetic factors of fiber traits in AD tetraploid cottons. [ABSTRACT FROM AUTHOR]

The use of non-invasive DNA-based sampling is becoming increasingly popular. However, the misidentification of individuals due to genotyping error (primarily allelic dropout) is a critical problem, especially when using individuals in the capture-mark-recapture (CMR) approach to estimate population size. We propose 2 simple and cost-effective tests, Examining Bimodality (EB) and Difference in Capture History (DCH), to determine whether a sample contains genotyping errors and the relative magnitude of the problem. These tests formalize currently used approaches for identifying genotyping errors. We evaluate the efficacy of these tests using simulated CMR data. Results show that both tests are effective at detecting genotyping errors and provide a strong indication of whether the data are error free. The EB and DCH tests apply to data in which multiple samples are associated with individuals, such as those generated by CMR sampling. Managers need to be able to identify and eliminate genotyping errors to produce population estimates that are both unbiased and scientifically defensible. [ABSTRACT FROM AUTHOR]

Capture-mark-recapture (CMR) estimates assume no misidentificafion of individuals captured and are extremely sensitive to identification errors. A large body of published literature has demonstrated that non-invasively derived genetic tags are error-prone, and the potential biases associated with these errors are large. We provided methods to reduce and evaluate these errors. Paetkau (2004, this issue), in his comments concerning our paper, argues that no formal, statistical error testing is necessary and that good laboratory practices are sufficient to remove all error. However, he provides only anecdotal evidence that this is the case. Given the presence of a variety of errors in genetic tags and the potential for large biases associated with these errors, we argue that scientific norms require formal tests to demonstrate tile absence of errors. The primary purpose of our study was to provide such tests in a manner that is not cost-prohibitive. [ABSTRACT FROM AUTHOR]

An introgression line derived from an interspecific cross between Oryza sativa and Oryza officinalis, IR54741-3-21-22 was found to be resistant to an Indian biotype of brown planthopper (BPH). Genetic analysis of 95 F3 progeny rows of a cross between the resistant line IR54741-3-21-22 and a BPH susceptible line revealed that resistance was controlled by a single dominant gene. A comprehensive RAPD analysis using 275 decamer primers revealed a low level of (7.1%) polymorphism between the parents. RAPD polymorphisms were either co-dominant (6.9%), dominant for resistant parental fragments (9.1%) or dominant for susceptible parental fragments (11.6%). Of the 19 co-dominant markers, one primer, OPA16, amplified a resistant parental band in the resistant bulk and a susceptible parental band in the susceptible bulk by bulked segregant analysis. RAPD analysis of individual F2 plants with the primer OPA16 showed marker-phenotype co-segregation for all, with only one recombinant being identified. The linkage between the RAPD marker OPA16938 and the BPH resistance gene was 0.52 cM in coupling phase. The 938 bp RAPD amplicon was cloned and used as a probe on 122 Cla I digested doubled haploid (DH) plants from a IR64xAzucena mapping population for RFLP inheritance analysis and was mapped onto rice chromosome 11. The OPA16938 RAPD marker could be used in a cost effective way for marker-assisted selection of BPH resistant rice genotypes in rice breeding programs. [ABSTRACT FROM AUTHOR]

• Oxidative titrations were applied to selectively quantify phenolic moieties in DOM. • Correlation between quantified phenolic DOM and electron donating capacity (EDC). • EDC of 27 DOM model compounds was determined. • Phenol concentration in DOM correlated to the formation of ClO 2 -byproducts. • Molecular tagging of phenols in DOM by formation of phenylacetates. The application of oxidants for disinfection or micropollutant abatement during drinking water and wastewater treatment is accompanied by oxidation of matrix components such as dissolved organic matter (DOM). To improve predictions of the efficiency of oxidation processes and the formation of oxidation products, methods to determine concentrations of oxidant-reactive phenolic, olefinic or amine-type DOM moieties are critical. Here, a novel selective oxidative titration approach is presented, which is based on reaction kinetics of oxidation reactions towards certain DOM moieties. Phenolic moieties were determined by oxidative titration with ClO 2 and O 3 for five DOM isolates and two secondary wastewater effluent samples. The determined concentrations of phenolic moieties correlated with the electron-donating capacity (EDC) and the formation of inorganic ClO 2 -byproducts (HOCl, ClO 2 −, ClO 3 −). ClO 2 -byproduct yields from phenol and DOM isolates and changes due to the application of molecular tagging for phenols revealed a better understanding of oxidant-reactive structures within DOM. Overall, oxidative titrations with ClO 2 and O 3 provide a novel and promising tool to quantify oxidant-reactive moieties in complex mixtures such as DOM and can be expanded to other matrices or oxidants. [Display omitted] [ABSTRACT FROM AUTHOR]

Separate genes conferring antibiotic drag resistance have been inserted into Fusarium graminearum and Fusarium moniliforme. These organisms are associated with stalk rot of corn, a disease of uncertain cause. Antibiotic resistant fungi were obtained by developing a gene transfer system using whole cells as recipients for DNA. Hygromycin B and benomyl-resistant colonies were isolated by treating fungal tissue with lithium acetate and adding plasmid vectors containing the respective genes which give drug resistance. The DNA was stably integrated into the fungal chromosome. Following plant inoculation, disease symptoms developed and the isolates were recovered on selective medium. In each case, these fungi retained the transformed phenotype, although extensive rearrangements and/or deletions occurred. Specific molecular tagging allows detailed studies of this interaction and should be of general use in situations involving complex multiple pathogen diseases. [ABSTRACT FROM AUTHOR]

Molecular characterization of Circulating Tumor Cells (CTCs) is still challenging, despite attempts to minimize the drawbacks of Whole Genome Amplification (WGA). In this paper, we propose a Next-Generation Sequencing (NGS) optimized protocol based on molecular tagging technology, in order to detect CTCs mutations while skipping the WGA step. MDA-MB-231 and MCF-7 cell lines, as well as leukocytes, were sorted into pools (2–5 cells) using a DEPArray™ system and were employed to set up the overall NGS procedure. A substantial reduction of reagent volume for the preparation of libraries was performed, in order to fit the limited DNA templates directly derived from cell lysates. Known variants in TP53, KRAS, and PIK3CA genes were detected in almost all the cell line pools (35/37 pools, 94.6%). No additional alterations, other than those which were expected, were found in all tested pools and no mutations were detected in leukocytes. The translational value of the optimized NGS workflow is confirmed by sequencing CTCs pools isolated from eight breast cancer patients and through the successful detection of variants. In conclusion, this study shows that the proposed NGS molecular tagging approach is technically feasible and, compared to traditional NGS approaches, has the advantage of filtering out the artifacts generated during library amplification, allowing for the reliable detection of mutations and, thus, making it highly promising for clinical use. [ABSTRACT FROM AUTHOR]

Genotyping error, often associated with low-quantity/quality DNA samples, is an important issue when using genetic tags to estimate abundance using capture-mark-recapture (CMR). dropout, an MS-Windows program, identifies both loci and samples that likely contain errors affecting CMR estimates. dropout uses a ‘bimodal test’, that enumerates the number of loci different between each pair of samples, and a ‘difference in capture history test’ (DCH) to determine those loci producing the most errors. Importantly, the DCH test allows one to determine that a data set is error-free. dropout has been evaluated in McKelvey & Schwartz (2004) and is now available online. [ABSTRACT FROM AUTHOR]

The tiger shrimp ( Penaeus monodon) is an important marine crustacean in terms of biological diversity and aquaculture resource. The shrimp is widespread across the Indo-Pacific region and shows apparent genetic differentiation among geographical populations. It is common practice to transport female brooders between different countries to seed the shrimp farms, posing potential problems of unwanted population admixture. We developed 23 polymorphic microsatellites for P. monodon (average HE = 0.936) and these microsatellites were applicable for studying population differentiation, identifying valid stocks and tagging nonindigenous farmed shrimps. [ABSTRACT FROM AUTHOR]