Organization and function of the phage Lambda chromosome

The process of prophage integration by phage λ and the function and structure of the chromosomal elements required for λ integration have been studied with the use of λ deletion mutants. Since att^φ, the substrate of the integration enzymes, is not essential for λ growth, and since att^φ resides in a portion of the λ chromosome which is not necessary for vegetative growth, viable λ deletion mutants were isolated and examined to dissect the structure of att^φ.

Deletion mutants were selected from wild type populations by treating the phage under conditions where phage are inactivated at a rate dependent on the DNA content of the particles. A number of deletion mutants were obtained in this way, and many of these mutants proved to have defects in integration. These defects were defined by analyzing the properties of Int-promoted recombination in these att mutants.

The types of mutants found and their properties indicated that att^φ has three components: a cross-over point which is bordered on either side by recognition elements whose sequence is specifically required for normal integration. The interactions of the recognition elements in Int-promoted recombination between att mutants was examined and proved to be quite complex. In general, however, it appears that the λ integration system can function with a diverse array of mutant att sites.

The structure of att^φ was examined by comparing the genetic properties of various att mutants with their location in the λ chromosome. To map these mutants, the techniques of heteroduplex DNA formation and electron microscopy were employed. It was found that integration cross-overs occur at only one point in att^φ and that the recognition sequences that direct the integration enzymes to their site of action are quite small, less than 2000 nucleotides each. Furthermore, no base pair homology was detected between att^φ and its bacterial analog, att^B. This result clearly demonstrates that λ integration can occur between chromosomes which have little, if any, homology. In this respect, λ integration is unique as a system of recombination since most forms of generalized recombination require extensive base pair homology.

An additional study on the genetic and physical distances in the left arm of the λ genome was described. Here, a large number of conditional lethal nonsense mutants were isolated and mapped, and a genetic map of the entire left arm, comprising a total of 18 genes, was constructed. Four of these genes were discovered in this study. A series of λdg transducing phages was mapped by heteroduplex electron microscopy and the relationship between physical and genetic distances in the left arm was determined. The results indicate that recombination frequency in the left arm is an accurate reflection of physical distances, and moreover, there do not appear to be any undiscovered genes in this segment of the genome.

Studies on the role of histones in the structure and function of chromatin

Studies on the dissociation of histones from chromatin by increasing concentrations of sodium deoxycholate (DOC) have shown that histrone II is removed at lowest concentrations of DOC, while slightly higher concentrations remove histones III and IV. Still higher concentrations remove histone I.

The complete separation of chromatin and ¹⁴C-DOC by sucrose sedimentation indicated that the binding of DOC to chromatin is readily and completely reversible.

The dissociation of histones from chromatin by increasing concentrations of related cholanic acids and some of their conjugated derivatives were studied. The results suggested that the driving force for the interaction between the cholanic acid anion and histones is the lowering of the activity coefficient of the cholanic acid anion which occurs when it is partially removed from solution by interaction with hydrophobic regions of the positively charged histones.

The role of histones in the structure of chromatin has been studied by comparing the effects of selective removal of histones from chromatin by increasing concentrations of DOC with those caused by NaCl (removes histone I at lowest concentrations, while higher concentrations remove histones II, III, and IV). Properties studied included thermal denaturation, sedimentation velocity, flow dichroism, relaxation times of molecules oriented in a flow field, and the irreversible disruption of a 130 S, cross-linked component of sheared chromatin. The data indicated that none of the structural or chemical parameters with which these properties are correlated show a dependence on the presence of one particular histone fraction.

The template activity (ability to prime a 0.2 M KC1 DNA-dependent RNA synthesis system catalyzed by E. coli RNA polymerase) increases from that of native chromatin (approximately 25 per cent of that pure DNA) to that of pure DNA in a fashion which shows a nearly linear relationship to the amount of histone coverage of the template. The precipitability of partially dehistonized chromatin samples in 0.15 M NaCl shows a large dependence on the presence of histone I.

Mass transfer from a cylinder to an air stream in axisymmetrical flow (an experimental study)

Mass transfer from wetted surfaces on one-inch cylinders with unwetted approach sections was studied experimentally by means of the evaporation of n-octane and n-heptane into an air stream in axisymmetrical flow, for Reynolds numbers from 5,000 to 310,000. A transition from the laminar to the turbulent boundary layer was observed to occur at Reynolds numbers from 10,000 to 15,000. The results were expressed in terms of the Sherwood number as a function of the Reynolds number, the Schmidt number, and the ratio of the unwetted approach length to the total length. Empirical formulas were obtained for both laminar and turbulent regimes. The rates of mass transfer obtained were higher than theoretical and experimental results obtained by previous investigators for mass and heat transfer from flat plates.

On the added mass of sphere in a circular cylinder considering real fluid effects

An experimental method combined with boundary layer theory is given for evaluating the added mass of a sphere moving along the axis of a circular cylinder filled with water or oil. The real fluid effects are separated from ideal fluid effects.

The experimental method consists essentially of a magnetic steel sphere propelled from rest by an electromagnetic coil in which the current is accurately controlled so that it only supplies force for a short time interval which is within the laminar flow regime of the fluid. The motion of the sphere as a function of time is recorded on single frame photographs using a short-arc multiple flash lamp with accurately controlled time intervals between flashes.

A concept of the effect of boundary layer displacement on the fluid flow around a sphere is introduced to evaluate the real fluid effects on the added mass. Surprisingly accurate agreement between experiment and theory is achieved.

Connectivity and Function of the Primate Insula

The insula is a mammalian cortical structure that has been implicated in a wide range of low- and high-level functions governing one’s sensory, emotional, and cognitive experiences. One particular role of this region is considered to be processing of olfactory stimuli. The ability to detect and evaluate odors has significant effects on an organism’s eating behavior and survival and, in case of humans, on complex decision making. Despite such importance of this function, the mechanism in which olfactory information is processed in the insula has not been thoroughly studied. Moreover, due to the structure’s close spatial relationship with the neighboring claustrum, it is not entirely clear whether the connectivity and olfactory functions attributed to the insula are truly those of the insula, rather than of the claustrum. My graduate work, consisting of two studies, seeks to help fill these gaps. In the first, the structural connectivity patterns of the insula and the claustrum in a non-human primate brain is assayed using an ultra-high-quality diffusion magnetic resonance image, and the results suggest dissociation of connectivity — and hence function — between the two structures. In the second study, a functional neuroimaging experiment investigates the insular activity during odor evaluation tasks in humans, and uncovers a potential spatial organization within the anterior portion of the insula for processing different aspects of odor characteristics.

Fast Lattice Green's Function Methods for Viscous Incompressible Flows on Unbounded Domains

In this thesis, a collection of novel numerical techniques culminating in a fast, parallel method for the direct numerical simulation of incompressible viscous flows around surfaces immersed in unbounded fluid domains is presented. At the core of all these techniques is the use of the fundamental solutions, or lattice Green’s functions, of discrete operators to solve inhomogeneous elliptic difference equations arising in the discretization of the three-dimensional incompressible Navier-Stokes equations on unbounded regular grids. In addition to automatically enforcing the natural free-space boundary conditions, these new lattice Green’s function techniques facilitate the implementation of robust staggered-Cartesian-grid flow solvers with efficient nodal distributions and fast multipole methods. The provable conservation and stability properties of the appropriately combined discretization and solution techniques ensure robust numerical solutions. Numerical experiments on thin vortex rings, low-aspect-ratio flat plates, and spheres are used verify the accuracy, physical fidelity, and computational efficiency of the present formulations.