Radial migration of DNA molecules in cylindrical flow. III. Circles and the effect of non-gaussian polymer statistics.

We have previously shown that DNA will migrate radially inward in a concentric-cylinder shear flow apparatus. We assumed gaussian chain statistics, and we considered only linear molecules. In this paper, we extend the analysis to closed circular molecules, and we consider non-gaussian statistics for both linears and circles. We find that, in good solvents, the inward radial migration velocity is more sensitive to the molecular weight than M5/2, which we previously reported for gaussian chains. Furthermore, linears migrate radially inward 8 times faster than do circles of the same molecular weight. This suggests the possibility of separating linear from circular DNA in solution.     

Mixture modeling for genome-wide localization of transcription factors

Chromatin immunoprecipitation followed by DNA microarray analysis (ChIP-chip methodology) is an efficient way of mapping genome-wide protein-DNA interactions. Data from tiling arrays encompass DNA-protein interaction measurements on thousands or millions of short oligonucleotides (probes) tiling a whole chromosome or genome. We propose a new model-based method for analyzing ChIP-chip data. The proposed model is motivated by the widely used two-component multinomial mixture model of de novo motif finding. It utilizes a hierarchical gamma mixture model of binding intensities while incorporating inherent spatial structure of the data. In this model, genomic regions belong to either one of the following two general groups: regions with a local protein-DNA interaction (peak) and regions lacking this interaction. Individual probes within a genomic region are allowed to have different localization rates accommodating different binding affinities. A novel feature of this model is the incorporation of a distribution for the peak size derived from the experimental design and parameters. This leads to the relaxation of the fixed peak size assumption that is commonly employed when computing a test statistic for these types of spatial data. Simulation studies and a real data application demonstrate good operating characteristics of the method including high sensitivity with small sample sizes when compared to available alternative methods.     

Automation of peak-tracking analysis of stepwise perturbed NMR spectra

We describe a new algorithmic approach able to automatically pick and track the NMR resonances of a large number of 2D NMR spectra acquired during a stepwise variation of a physical parameter. The method has been named Trace in Track (TinT), referring to the idea that a gaussian decomposition traces peaks within the tracks recognised through 3D mathematical morphology. It is capable of determining the evolution of the chemical shifts, intensity and linewidths of each tracked peak.The performances obtained in term of track reconstruction and correct assignment on realistic synthetic spectra were high above 90% when a noise level similar to that of experimental data were considered. TinT was applied successfully to several protein systems during a temperature ramp in isotope exchange experiments. A comparison with a state-of-the-art algorithm showed promising results for great numbers of spectra and low signal to noise ratios, when the graduality of the perturbation is appropriate. TinT can be applied to different kinds of high throughput chemical shift mapping experiments, with quasi-continuous variations, in which a quantitative automated recognition is crucial.     

Bayesian robust inference for differential gene expression in microarrays with multiple samples

We consider the problem of identifying differentially expressed genes under different conditions using gene expression microarrays. Because of the many steps involved in the experimental process, from hybridization to image analysis, cDNA microarray data often contain outliers. For example, an outlying data value could occur because of scratches or dust on the surface, imperfections in the glass, or imperfections in the array production. We develop a robust Bayesian hierarchical model for testing for differential expression. Errors are modeled explicitly using a t-distribution, which accounts for outliers. The model includes an exchangeable prior for the variances, which allows different variances for the genes but still shrinks extreme empirical variances. Our model can be used for testing for differentially expressed genes among multiple samples, and it can distinguish between the different possible patterns of differential expression when there are three or more samples. Parameter estimation is carried out using a novel version of Markov chain Monte Carlo that is appropriate when the model puts mass on subspaces of the full parameter space. The method is illustrated using two publicly available gene expression data sets. We compare our method to six other baseline and commonly used techniques, namely the t-test, the Bonferroni-adjusted t-test, significance analysis of microarrays (SAM), Efron's empirical Bayes, and EBarrays in both its lognormal-normal and gamma-gamma forms. In an experiment with HIV data, our method performed better than these alternatives, on the basis of between-replicate agreement and disagreement.     

The resolution of aneuploid DNA stem lines by flow cytometry: limitations imposed by the coefficient of variation and the percentage of aneuploid nuclei

Factors important in the resolution of cell sub-populations with differing DNA contents were investigated using an EPICS C flow cytometer. Software is available for the EPICS C which permits data from any two histograms to be superimposed or added together before display. Samples of fresh and archival thyroid tissue, stained with propidium iodide, were analysed on the flow cytometer and the peak channel number noted. The photomultiplier (PMT) voltage was increased and the sample analysed again producing a second histogram with a higher peak channel number. The two histograms were added together to simulate a cell suspension with two sub-populations with a different DNA content. By systematically altering the PMT voltage and the number of nuclei included in each analysis, it was possible to examine the importance of DNA index and the percentage of tumor cells with an aneuploid DNA content for both fresh and paraffin-embedded thyroid nuclei. The crucial importance of achieving a low coefficient of variation (CV) was demonstrated and consequently the reservations that pertain when archival material is studied, particularly in tumours where DNA aneuploidy is frequently expressed with a low DNA index.     

Bivariate flow karyotyping of human chromosomes: evaluation of variation in Hoechst 33258 fluorescence, chromomycin A3 fluorescence, and relative chromosomal DNA content.

The total variation of chromosome peak positions, in bivariate distributions of Hoechst 33258 and chromomycin A3 fluorescence of 19 healthy individuals, was compared with the experimental variation, determined from 23 bivariate distributions of chromosomes prepared separately from a single cell lineage. The experimental variation in Hoechst and chromomycin fluorescence and the relative chromosomal DNA content were determined from experiments performed over several days. The additional variance contributed by time was the same as the daily variance. The accuracy by which the relative chromosomal DNA content can be calculated from bivariate peak positions was investigated. A least squares method was used to fit the distributions of relative DNA content, obtained, respectively, from mono- and bivariate flow analyses of chromosomes from the same cell lineage. In general the DNA contents match quite well, but for a few chromosomes a difference was found, statistically discernible at the 5% level. The average relative chromosomal DNA content of the chromosomes from the 19 normal individuals, calculated from bivariate peak positions, showed a linear relation with the estimates published by other investigators.     

Analysis of cellular DNA distributions

An analytical formula for calculating peak channel ratios in fluorescent cytophotometric determinations of DNA content per cell was derived to assess the effects of inaccuracies in the model-dependent derivation of S-phase cell populations and of systematic instrumental errors. The DNA distribution histograms usually have two peaks, corresponding to the 2C DNA content of G₁ cells and to the 4C DNA content of G₂ and M cells. In the presence of S-phase cells, the ratio of peak channels G₂/G₁ becomes less than 2. The calculation uses the model-dependent number of S-phase cells per channel and instrumental resolution to obtain G₂/G₁. The peak channel ratio calculated in this way decreases with increasing coefficient of variation and increasing proportion of S-phase cells. The calculated G₂/G₁ peak channel ratios were compared with 17 experimental values ranging from 1.68 to 2.08. Significant differences were found for two experiments, and the calculated G₂/G₁ ratios were systematically low by ≈4% for the other experiments. When this systematic difference in predicted peak channel ratios is taken into account, the formula predicts the observed ratios with an accuracy of 1% showing the dominant effect of S-phase cells in modifying the observed spectrum. The possible experimental effects leading to the observed systematic discrepancy are discussed A programmable pocket calculator program to perform these calculations is also described in detail.     

Comparison of the thermal denaturation behaviour of DNA-solutions and metaphase chromosome preparations in suspension.

Hyperchromicity measurements are well established to analyse the thermal denaturation behaviour of pure DNA sequences in solution. Here, we show that under appropriate experimental conditions this technique can also be applied to study thermally controlled conformation changes of higher order DNA-protein complexes as for instance metaphase chromosome preparations in suspension. A computer controlled sensitive, upright double beam photometer with a heatable cuvette was constructed. Measurements of the temperature dependent extinction of both, solutions and particle suspensions are possible, since sedimentation effects of particles can be neglected due to the vertical optical axis in the probe cuvette. Thermal denaturation of metaphase chromosome preparations of human and Chinese hamster cells was investigated and compared to melting profiles of DNA solutions for two excitation wavelengths, 256 and 313 nm. The influence of neutral and low pH was considered. The results indicate that metaphase chromosome preparations show a thermal denaturation behaviour different from pure DNA. Whereas DNA solutions showed one pH dependent melting peak at 256 nm only, the peak pattern of metaphase chromosome preparations showed a large variability both at 256 and 313 nm. At neutral pH, in two temperature regions (40-55 degrees C and 75-82 degrees C) peaks were found indicating chromosome typical conformation changes independently from the mammalian cell species (Chinese hamster, human). In contrast to pure DNA, no typical reduction in the temperatures of peak maxima with decreasing pH was found for metaphase chromosome preparations of both cell types. These results may be relevant for further systematic studies of efficient thermal probe/target denaturation procedures in non enzymatic DNA-chromosome in situ hybridisation.     

Coding DNA sequences: statistical distributions

The base distributions in coding DNA sequences (CDS) are investigated. We explore the scaling properties of the 4-dimensional directed random walk and compare them with that for the DNA sequences. Inference from these observation are, however, contradicted by alternate analysis using factorial moments. To resolve this conflict we look directly at the nucleotide base distributions. In all the cases the base distributions change from gaussian to non-gaussian as the scale size is increased. The CDS, therefore, have nucleotide distributions different from the random.     

STUDIES ON DNA FROM NORMAL AND SCRAPIE-AFFECTED MOUSE BRAIN

Abstract— DNA has been extracted from normal and scrapie affected mouse brain fractions, 48 h after the injection of [³H]thymidine precursors. The extracted DNA has been subjected to fractionation on hydroxyapatite columns and CsCl gradients. The specific activity of double stranded nuclear DNA is two to three times higher when extracted from scrapie-affected brain than from normal brain, but there is no apparent difference in the number of counts associated with double stranded mitochondrial DNA extracted from similar numbers of normal and scrapie affected brains. DNA from the large granule fraction of scrapie affected brain contains a peak of counts, melting off hydroxyapatite columns before the double stranded peak, consistent with the presence in scrapie brain of trace amounts of a small single stranded DNA.     

The perception of a dotted line in noise: a model of good continuation and some experimental results

A formal model is proposed, describing how the perceptual interpretation of dot figures is guided by the Gestalt rule of good continuation. The algorithm will be restricted to figures with a collinear dot array (line) embedded in a background of randomly placed dots. The model, CODE-2, is an elaboration of the model, CODE-1, of grouping dots on the basis of the Gestalt rule of (relative) proximity, and consists of the introduction of non-circular symmetric gaussian distribution functions for the representation of the orientation dependent strength of interaction between collinear dots. Supra-threshold contours of the function, resulting from a superposition on each dot of the gaussian functions, are assumed to predict the perceptual grouping of the dots. A quantitative measure for the perceptual salience of dotted lines was defined as the contrast between the internal coherence of the line dots, and their interference with the noise dots. For 20 stimuli the CODE-2 grouping of the dots is reported, together with the results of a line-in-noise latency experiment. There was a significant correlation between the predicted saliences and the experimental results. The results support the usefulness of representing good continuation between collinear dots by non-circular symmetric gaussian distribution functions.     

Geometric and many-particle aspects of transmitter binding.

We investigate the various reactivity patterns possible when several transmitter molecules, released at one side of a synaptic gap, diffuse and bind reversibly to a single receptor at the other end. In the framework of a one-dimensional approximation, the complete time, reactivity, concentration and gap-width dependence are determined, using a rigorous theoretical and computational approach to the many-body aspects of this problem. The time dependence of the survival probability is found to consist of up to four phases. These include a short delay followed by gaussian, power-law, and exponential decay phases. A rigorous expression is derived for the long-time exponent and approximate expressions are obtained for describing the short-time gaussian phase.     

Analysis of cellular DNA distributions. I. G2/G1 channel ratios.

In the influencent-flow cytophotometric measurement of cellular DNA content the DNA distributions usually have two peaks. The second peak, which corresponds to the 4C DNA content of G2 and M cells, is often positioned at lower values of DNA content than twice that of the 2C DNA peak which contains G1 cells. Computerized numerical analyses were performed on artificial DNA distributions in which the proportion of S-phase cells was varied. It was demonstrated that the contribution of late S-phase cells to the 4C DNA peak in the histogram shifts the second peak to a position below twice the 2C DNA value. Also, increasing the coefficient of variation of the DNA measurement shifts the second peak position to lower values. A group of 33 DNA distribution histograms we found to have an average G2/G1 peak position ratio of 1.90, in keeping with typical values obtained from the numerical analysis of the artificial populations.     

Analysis of cellular DNA distributions

In the fluorescent-flow cytophotometric measurement of cellular DNA content the DNA distributions usually have two peaks. The second peak, which corresponds to the 4C DNA content of G2 and M cells, is often positioned at lower values of DNA content than twice that of the 2C DNA peak which contains G1 cells. Computerized numerical analyses were performed on artificial DNA distributions in which the proportion of S-phase cells was varied. It was demonstrated that the contribution of late S-phase cells to the 4C DNA peak in the histogram shifts the second peak to a position below twice the 2C DNA value. Also, increasing the coefficient of variation of the DNA measurement shifts the second peak position to lower values. A group of 33 DNA distribution histograms was found to have an average G2/G1 peak position ratio of 1.90, in keeping with typical values obtained from the numerical analysis of the artificial populations.     

Radial migration of DNA molecules in cylindrical flow. II. The non-draining model and possible application to fractionation

The radial-migration calculation of the preceding paper is extended to the non-draining polymer. The results show an inward radial velocity that depends on the 5/2 power of the molecular weight, assuming gaussian statistics for the chain. This sensitivity to size serves as the basis for a method of separating DNA molecules in solution according to their size. Normal stress (Weissenberg) effects are also discussed.     

A whole-genome analysis using robust asymmetric distributions

This study is aimed at improving the analysis of data used in identifying marker-associated effects on quantitative traits, specifically to account for possible departures from a Gaussian distribution of the trait data and to allow for asymmetry of marker effects attributable to phenotypic divergence between parental lines. A Bayesian procedure for analysing marker effects at the whole-genome level is presented. The procedure adopts a skewed t-distribution as a prior distribution of marker effects. The model with the skewed t-process includes Gaussian prior distributions, skewed Gaussian prior distributions and symmetric t-distributions as special cases. A Markov Chain Monte Carlo algorithm for obtaining marginal posterior distributions of the unknowns is also presented. The method was applied to a dataset on three traits (live weight, carcass length and backfat depth) measured in an F2 cross between Iberian and Landrace pigs. The distribution of marker effects was clearly asymmetric for carcass length and backfat depth, whereas it was symmetric for live weight. The t-distribution seems more appropriate for describing the distribution of marker effects on backfat depth.     

A Stochastic Epidemic Model with Seasonal Variations in Infection Rate

In this paper we consider a modification of Bailey's stochastic model for the spread of an epidemic when there are seasonal variations in infection rate. The resulting nonlinear model is analyzed by employing the diffusion approximation technique. We have shown that for a large population the process, on suitable scaling and normalization, converges to a non-stationary Ornstein-Uhlenbeck process. Consequently the number of infectives has in the steady state a gaussian distribution.     

N^7＋重离子注入和贯穿后小麦种胚的期外DNA合成效应

用＾３ＨＴｄＲ掺入法研究了经Ｎ＾７＋重离子注入贯穿处理的８２５７９小麦和８８１２小麦种胚的ＤＮＡ合成动态。结果发现,未经Ｎ＾７＋重离子任何处理的两个小麦品种的对照种胚,在萌发早期（２０ｈ内）仅存在一个ＤＮＡ合成峰（于萌发的第１４ｈ）,而经过Ｎ＾７＋重离子注入和贯穿处理的小麦种胚则存在两个ＤＮＡ合成峰（分别于萌发的８－１０ｈ和１４－１６ｈ）,该种子经ＤＮＡ修复合成的抑制剂咖啡因处理后,第一个ＤＮＡ合