Initiation of V(D)J recombination in a cell-free system

Cells performing V(D)J recombination make specific cuts in DNA at recombination signal sequences. Here, we show that nuclear extracts of pre-B cell lines carry out this specific cleavage. The products of cleavage are the same as found previously in thymocytes: full-length, blunt, 5'-phosphorylated signal ends, and covalently sealed (hairpin) coding ends. A complete signal sequence is required. Recombinant RAG1 protein greatly increases activity and complements an inactive extract from a RAG1 (-/-) pre-B cell line. When the extracts are fractionated, cleavage activity correlates with the presence of RAG2 protein. These results suggest that RAG1 and RAG2 are components of the V(D)J recombinase

A visual display enhancing comfort by counteracting airsickness

A simulator study has been conducted demonstrating a positive effect on airsickness by utilizing a 3D artificial Earth-fixed visual pattern. Participants were exposed to the same turbulent physical aircraft motion in a simulator three times in a row, each time using a different visual cue. In one condition only the interior of the simulator cabin was visible. In another condition an Earth-fixed star field moving opposite the simulator cabin was projected in front of the participant. In a third condition the same star field was used, however with additional anticipatory information by means of a rollercoaster like track showing the trajectory to go. Participants were asked for their misery and joyfulness ratings at fixed time instants using an 11-points misery scale (no problems-vomiting) and joyfulness scale (unpleasant-pleasant). The results showed that viewing an Earth-fixed visual frame moving instantaneously opposite the cabin motion did reduce motion sickness significantly by a factor of 1.6, thereby improving comfort. This condition could be applied in air transport, where often a monitor is available in the back of the seat ahead. The largest effect, i.e., a reduction by a factor of 4.2 was realized by adding anticipatory information. Although it is not possible to predict the effect of turbulence on the aircraft motion yet, an anticipatory display might already be applicable in other domains, such as at sea by using a wave radar and a ship motion model. © 2011 Elsevier B.V. All rights reserved

Quantification of viable eggs of the potato cyst nematodes (Globodera spp.) using either trehalose or RNA-specific Real-Time PCR

Two novel methods for the quantitative estimation of the number of viable eggs of the potato cyst nematodes (Globodera pallida and G. rostochiensis) were tested and compared with visual inspection. One is based on the loss of membrane integrity upon death and uses trehalose (a disaccharide) as a marker, the second test exploits the rapid degeneration of mRNA upon decease with a RNA-specific Real-Time Polymerase Chain Reaction (RT-PCR) assay. The viability of eggs in suspensions with different numbers of eggs was determined morphologically and was compared with both trehalose and elongation-factor-1-alpha (EF1a) mRNA measurements. The trehalose assay provided results that were close to those of the visual assessment using a microscope but only when samples contained low numbers of eggs. The lowest detectable value is 1.1 egg in the original sample and small differences in the number of viable eggs can be detected. Unfortunately, trehalose measurements reached a saturation limit at 1 cyst 10 µl-1; therefore, samples with nematode numbers above 262 eggs have to be diluted. The presence of dead cysts did not have a negative effect on the trehalose measurements. However, the use of egg suspensions instead of encysted eggs improved both the trehalose absorbance and the reliability of the measurements. When cysts were exposed to a treatment with allylisothiocyanate, the trehalose measurement detected the presence of more viable eggs than a hatching assay. The RT-PCR assay required a minimum of 30 eggs before detection occurred but can detect up to 8000 eggs in a 25 µl sample, which is an advantage when samples with high PCN infestations have to be processed. However, the confidence intervals (CI) of the RT-PCR assay are larger than those of the trehalose assay, which results in a high variation of single measurements. For example, at a density of 210 eggs in the original sample the 95% CI for the trehalose assay covers 191-228 eggs, and the 95% CI for the RT-PCR assay for G. pallida lies between 73 and 602 eggs and for G. rostochiensis between 59 and 745 eggs. Trials with field samples using both methods supported the laboratory tests. 95% of the field samples tested with the trehalose assay lie within the CI of the standard curve compared to 58% of the RT-PCR tested samples for G. pallida. The measurements of the field samples of G. pallida and G. rostochiensis populations using both methods resulted in larger numbers of viable eggs being detected compared to a hatching test. Neither of the investigated methods in their current state of development is optimal for use as a substitute for the visual inspection used in monitoring labs. The variance of the RT-PCR assay is too high if used for quantitative monitoring; the density range of eggs that can be detected using the trehalose assay is too small

Quantification of viable eggs of the potato cyst nematodes (Globodera spp.) using either trehalose or RNA-specific Real-Time PCR

Two novel methods for the quantitative estimation of the number of viable eggs of the potato cyst nematodes (Globodera pallida and G. rostochiensis) were tested and compared with visual inspection. One is based on the loss of membrane integrity upon death and uses trehalose (a disaccharide) as a marker, the second test exploits the rapid degeneration of mRNA upon decease with a RNA-specific Real-Time Polymerase Chain Reaction (RT-PCR) assay. The viability of eggs in suspensions with different numbers of eggs was determined morphologically and was compared with both trehalose and elongation-factor-1-alpha (EF1a) mRNA measurements. The trehalose assay provided results that were close to those of the visual assessment using a microscope but only when samples contained low numbers of eggs. The lowest detectable value is 1.1 egg in the original sample and small differences in the number of viable eggs can be detected. Unfortunately, trehalose measurements reached a saturation limit at 1 cyst 10 µl-1; therefore, samples with nematode numbers above 262 eggs have to be diluted. The presence of dead cysts did not have a negative effect on the trehalose measurements. However, the use of egg suspensions instead of encysted eggs improved both the trehalose absorbance and the reliability of the measurements. When cysts were exposed to a treatment with allylisothiocyanate, the trehalose measurement detected the presence of more viable eggs than a hatching assay. The RT-PCR assay required a minimum of 30 eggs before detection occurred but can detect up to 8000 eggs in a 25 µl sample, which is an advantage when samples with high PCN infestations have to be processed. However, the confidence intervals (CI) of the RT-PCR assay are larger than those of the trehalose assay, which results in a high variation of single measurements. For example, at a density of 210 eggs in the original sample the 95% CI for the trehalose assay covers 191-228 eggs, and the 95% CI for the RT-PCR assay for G. pallida lies between 73 and 602 eggs and for G. rostochiensis between 59 and 745 eggs. Trials with field samples using both methods supported the laboratory tests. 95% of the field samples tested with the trehalose assay lie within the CI of the standard curve compared to 58% of the RT-PCR tested samples for G. pallida. The measurements of the field samples of G. pallida and G. rostochiensis populations using both methods resulted in larger numbers of viable eggs being detected compared to a hatching test. Neither of the investigated methods in their current state of development is optimal for use as a substitute for the visual inspection used in monitoring labs. The variance of the RT-PCR assay is too high if used for quantitative monitoring; the density range of eggs that can be detected using the trehalose assay is too small

Phase transition properties of clustered travelling salesman problem instances generated with evolutionary computation

This paper introduces a generator that creates problem instances for the Euclidean symmetric travelling salesman problem. To fit real world problems, we look at maps consisting of clustered nodes. Uniform random sampling methods do not result in maps where the nodes are spread out to form identifiable clusters. To improve upon this, we propose an evolutionary algorithm that uses the layout of nodes on a map as its genotype. By optimising the spread until a set of constraints is satisfied, we are able to produce better clustered maps, in a more robust way. When varying the number of clusters in these maps and, when solving the Euclidean symmetric travelling salesman person using Chained Lin-Kernighan, we observe a phase transition in the form of an easy-hard-easy patter