Efficient random subcloning of DNA sheared in a recirculating point-sink flow system

Based on a high-performance liquid chromatographic pump, we have built a device that allows recirculation of DNA through a 63-microm orifice with ensuing fractionation to a minimum fragment size of approximately 300 base pairs. Residence time of the DNA fragments in the converging flow created by a sudden contraction was found to be sufficiently long to allow extension of the DNA molecules into a highly extended conformation and, hence, breakage to occur at midpoint. In most instances, 30 passages sufficed to obtain a narrow size distribution, with >90% of the fragments lying within a 2-fold size distribution. The shear rate required to achieve breakage was found to be inversely proportional to the 1.0 power of the molecular weight. Compared with a restriction digest, up to 40% of all fragments could be cloned directly, with only marginal improvements in cloning efficiency having been observed upon prior end repair with Klenow, T4 polymerase or T4 polynucleotide kinase. Sequencing revealed a fairly random distribution of the fragments.     

The band areas of proteins determined by fluorescent scanning in the commercial automated gel electrophoresis apparatus

An automated gel electrophoresis apparatus, recently available commercially, allows one to follow the band during electrophoresis in real time, and lends itself therefore to an evaluation of bandwidth as a function of migration time (the dispersion coefficient), resolution and band shape. These determinations assume the constancy of band area with migration time and at various gel concentrations. The purpose of the present study was to verify these assumptions. Representative proteins and sodium dodecyl sulfate (SDS)-proteins, either natively fluorescent or fluorescein carboxylate labeled, were found to exhibit band areas which approach constancy as a function of migration time in both agarose and polyacrylamide gel electrophoresis, provided that (i) the protein concentration under the band was low enough to obviate self-quenching of fluorescence; (ii) the separation of the protein of interest from contaminants had progressed sufficiently during the time at which band areas were measured; (iii) the baseline under the peak was sufficiently well defined. However, band areas decrease with increasing gel concentration. Protein peaks exhibited leading and trailing tails. The ratio of the combined tail area to total area appeared to be near-constant at varying migration times. However, that ratio increases with increasing gel concentration. The tail area does not appear to be an artifact of fluorometric detection since it is reproduced upon fluorimetric analysis of the protein eluted from gel slices after electrophoresis. However, it may be due to photochemical destruction under the conditions of repetitive fluorometric peak detection.     

IMPACCT: Methodology and Tools for Power-Aware Embedded Systems

Power-aware systems are those that must exploit a widerange of power/performance trade-offs in order to adapt to the power availabilityand application requirements. They require the integration of many novel powermanagement techniques, ranging from voltage scaling to subsystem shutdown.However, those techniques do not always compose synergistically with eachother; in fact, they can combine subtractively and often yield counterintuitive,and sometimes incorrect, results in the context of a complete system. Thiscan become a serious problem as more of these power aware systems are beingdeployed in mission critical applications.To address the problem of technique integration for power-aware embedded systems, we propose a new design tool framework called IMPACCT and the associated design methodology. The system modeling methodology includes application model for capturing timing/powerconstraints and mode dependencies at the system level. The tool performs power-awarescheduling and mode selection to ensure that all timing/power constraintsare satisfied and that all overhead is taken into account. IMPACCT then synthesizesthe implementation targeting a symmetric multiprocessor platform. Experimentalresults show that the increased dynamic range of power/performance settingsenabled a Mars rover to achieve significant acceleration while using lessenergy. More importantly, our tool correctly combines the state-of-the-arttechniques at the system level, thereby saving even experienced designersfrom many pitfalls of system-level power management.     

The effect of mischmetal addition on the structure and mechanical properties of a cast Al-7Si-0.3Mg alloy containing excess iron (up to 0.6 Pct)

The effect of Fe content (0.2 to 0.6 pct) on the microstructure and mechanical properties of a cast Al-7Si-0.3Mg (LM 25/356) alloy has been investigated. Further, 1 pct mischmetal (MM) additions (a mixture of rare-earth (RE) elements) were made to these alloys, and their mechanical properties at room and at elevated temperatures (up to 200 °C) were evaluated. A structure-property correlation on this alloy was attempted using optical microstructure analysis, fractographs, X-ray diffraction, energy-dispersive analysis of X-rays (EDX), and quantitative metallography by image analysis. An increase in Fe content increased the volume percentage of Fe-bearing intermetallic compounds (β and π phases), contributing to the loweryield strength (YS), ultimate tensile strength (UTS), percentage elongation, and higher hardness. An addition of 1 pct MM to the alloys containing 0.2 and 0.6 pct Fe was found to refine the microstructure; modify the eutectic silicon and La, Ce, and Nd present in the MM; form different intermetallic compounds with Al, Si, Fe, and Mg; and improve the mechanical properties of the alloys both at room and elevated temperatures.     

Pedestrian detection and tracking at crossroads

This paper presents a system that can perform pedestrian detection and tracking using vision-based techniques. A very important issue in the field of intelligent transportation system is to prevent pedestrians from being hit by vehicles. Recently, a great number of vision-based techniques have been proposed for this purpose. In this paper, we propose a vision-based method, which combines the use of a pedestrian model as well as the walking rhythm of pedestrians to detect and track walking pedestrians. Through integrating some spatial and temporal information grabbed by a vision system, we are able to develop a reliable system that can be used to prevent traffic accidents happened at crossroads. In addition, the proposed system can deal with the occlusion problem. Experimental results obtained by executing some real world cases have demonstrated that the proposed system is indeed superb.     

Effects of cocaine on dopamine receptor gene expression: a study in the postmortem human brain

The effects of chronic cocaine exposure on dopamine D1 and D2 receptor gene expression in the human brain were studied in postmortem samples from chronic cocaine abusing and matched control subjects. Using in situ hybridization of receptor autoradiography to examine messenger ribonucleic acid (RNA) and binding sites, respectively, neither D1 nor D2 receptor expression was found to be changed in the nucleus accumbens, caudate, putamen, or substantia nigra of the cocaine-exposed subjects. Although chronic cocaine exposure can produce alterations in dopaminergic neurotransmission, sustained compensatory changes in dopamine receptor expression do not appear to occur in the human.     

Symptom correlates of attentional improvement following hospitalization for a first episode of affective psychosis

BACKGROUND: This study examined patients with a first-episode of affective psychosis during acute and compensated states in order to determine whether changes in attentional functioning over time were accompanied by changes in the severity of psychotic or affective symptoms. METHODS: Attentional performance was measured in patients (n = 27) using the degraded-stimulus continuous Performance Test (CPT) and symptoms were assessed at the time of index hospitalization, and 2 months after discharge. A comparison group of normal volunteers (n = 31) also performed the CPT two months apart. RESULTS: Patients performed significantly worse than controls at the initial testing but not at follow-up. The improvement in attentional performance significantly correlated with decreased severity of manic symptoms. CONCLUSIONS: Results suggest attentional dysfunction is a state-dependent characteristic of mania, and may provide an additional measure of clinical improvement following treatment.     

Carcinogenic polycyclic aromatic hydrocarbons increase intracellular Ca2+ and cell proliferation in primary human mammary epithelial cells

A method was developed to determine in eggs 2 components [4,6-dimethyl-2-hydroxypyrimidine and 1,3-bis(4-nitrophenyl)urea] of the anticoccidial drug nicarbazin, used to treat poultry. Samples were extracted with acetonitrile, and the extracts were washed with hexane and evaporated to dryness before analysis by liquid chromatography/mass spectrometry with atmospheric pressure chemical ionization. By switching from positive to negative ion monitoring and using gradient elution, both components were measured within one run. The limit of quantitation of the assay was 10 ng/g for each component. The results of a preliminary feeding trial in which chickens were fed contamination levels of the drug are also reported.