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 structure and management of service level agreements innetworks

The paper proposes a structure for quality-of-service (QoS)-centered service level agreements (SLA), and a framework for their real-time management in multiservice packet networks. The SLA is structured to be fair to both parties, the service provider and their customer. The SLA considered here are for QoS assured delivery of aggregate bandwidth from ingress to egress nodes; however, the control and signaling is for the more granular flows or calls. A SLA monitoring scheme is presented in which revenue is generated by the admission of flows into the network, and penalty incurred when flows are lost in periods when the service provider is not SLA compliant. In the SLA management scheme proposed, the results of a prior off-line design are used, in conjunction with measurements taken locally at ingress nodes, to classify the loading status of routes. The routing and resource management are based on virtual partitioning and its supporting mechanism of bandwidth protection. The effectiveness of SLA management is measured by the robustness in performance in the presence of substantial diversity in actual traffic conditions. A simulation testbed called D'ARTAGNAN has been built from which we report numerical results for a case study. The results show that the SLA management scheme is robust, fair and efficient over a broad range of traffic conditions     

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.     

An artificial neural network approach on parametric optimization of laser micro-machining of die-steel

In the present research, laser micro machining (LMM) of tungsten-molybdenum general purpose high speed steel (Rex M2) has been studied. Selection of optimum machining parameter combinations for obtaining higher depth of groove and smaller height of recast layer is a challenging task in LMM due to the presence of a large number of process variables. There is no perfect combination of parameters which can simultaneously result in both the highest depth of groove and lowest height of recast layer. This paper presents an attempt to develop a strategy for predicting the optimum machining parameter setting for the generation of the maximum depth of groove with minimum height of recast layer. A feed forward back-propagation neural network has been developed to model the machining process. The model, after proper training, is capable of predicting the response parameters as a function of four different control parameters. Experimental results demonstrate that the machining model is suitable and the optimization strategy satisfies practical requirements. The developed model has been found to be quite unique, powerful and flexible.     

Modified CMA based blind multiuser equaliser with decision directed scheme

A modified constant modulus (CM) algorithm based blind multiuser equaliser with a decision directed (DD) scheme is developed that does not require any mutual decorrelation and is suitable for downlink direct sequence code division multiple access systems. The performance of the proposed time domain adaptive equaliser is evaluated with respect to usual parameters and is found to perform better than the concurrent CM algorithm and DD scheme.     

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.