A randomized library approach to identifying functional lox site domains for the Cre recombinase

The bacteriophage P1 Cre/loxP site-specific recombination system is a useful tool in a number of genetic engineering processes. The Cre recombinase has been shown to act on DNA sequences that vary considerably from that of its bacteriophage recognition sequence, loxP. However, little is known about the sequence requirements for functional lox-like sequences. In this study, we have implemented a randomized library approach to identify the sequence characteristics of functional lox site domains. We created a randomized spacer library and a randomized arm library, and then tested them for recombination in vivo and in vitro. Results from the spacer library show that, while there is great plasticity, identity between spacer pairs is the most important factor influencing function, especially in in vitro reactions. The presence of one completely randomized arm in a functional loxP recombination reaction revealed that only three wild-type loxP arms are necessary for successful recombination in Cre-expressing bacteria, and that there are nucleotide preferences at the first three and last three positions of the randomized arm for the most efficiently recombined sequences. Finally, we found that in vitro Cre recombination reactions are much more stringent for evaluating which sequences can support efficient recombination compared to the 294-CRE system

Hydrodynamics of membranes: the bilayer aspect and adhesion

We review recent work on the dynamics of membranes. First, we discuss the viscous modes of a free bilayer membrane, taking into account the coupling between bending and the local density of the two monolayers. Apart from important corrections to the conventional bending mode, we obtain a second slow mode which is essentially a fluctuation in the density difference of the two monolayers, damped by intermonolayer friction. Estimates for a stack of membranes show reasonable agreement with a recent spin-echo study of membrane undulations. Second, we discuss the dispersion relation for both fluid and polymerized membranes bound to a substrate by an attractive potential. The interplay between the hydrodynamic damping by the surrounding liquid and the restoring force in the binding potential leads to a q2 dependence of the damping rate γ at small wavevector q. With increasing q, various crossovers can occur, including the possibility of nonmonotonic damping in which γ decreases with q as ∼ 1/q

Viscous modes of fluid bilayer membranes

We determine the dispersion relation for a fluid bilayer membrane, taking into account the coupling between bending and the local density of the two monolayers. Apart from important corrections to the conventional bending mode, we obtain a second slow mode which is essentially a fluctuation in the density difference of the two monolayers, damped by inter-monolayer friction. Estimates for a stack of membranes show reasonable agreement with a recent spin-echo study of membrane undulations

Effective Temperatures of a Driven System Near Jamming

Fluctuations in a model of a sheared, zero-temperature foam are studied numerically. Five different quantities that reduce to the true temperature in an equilibrium thermal system are calculated. All five have the same shear-rate dependence, and three have the same value. Near the onset of jamming, the relaxation time is the same function of these three temperatures in the sheared system as of the true temperature in an unsheared system. These results imply that statistical mechanics is useful for the system and provide strong support for the concept of jamming.Comment: 4 pages, 4 postscript figure

The Melting of Aqueous Foams

Diffusing-wave spectroscopy measurements show that ordinarily solid aqueous foams flow by a series of stick-slip avalanche-like rearrangements of neighboring bubbles from one tight packing configuration to another. Contrary to a recent prediction, the distribution of avalanche sizes do not obey a power-law distribution characteristic of self-organized criticality. This can be understood from a simple model of foam mechanics based on bubble-bubble interactions

Statistics of Shear-Induced Rearrangements in a Two-Dimensional Model Foam

Under steady shear, a foam relaxes stress through intermittent rearrangements of bubbles accompanied by sudden drops in the stored elastic energy. We use a simple model of foam that incorporates both elasticity and dissipation to study the statistics of bubble rearrangements in terms of energy drops, the number of nearest neighbor changes, and the rate of neighbor-switching (T1) events. We do this for a two-dimensional system as a function of system size, shear rate, dissipation mechanism, and gas area fraction. We find that for dry foams, there is a well-defined quasistatic limit at low shear rates where localized rearrangements occur at a constant rate per unit strain, independent of both system size and dissipation mechanism. These results are in good qualitative agreement with experiments on two-dimensional and three-dimensional foams. In contrast, we find for progessively wetter foams that the event size distribution broadens into a power law that is cut off only by system size. This is consistent with criticality at the melting transition

Hubble Space Telescope WFPC-2 Imaging of Cassiopeia A

The young SNR Cassiopeia A was imaged with WFPC-2 through four filters selected to capture the complete velocity range of the remnant's main shell in several important emission lines. Primary lines detected were [O III] 4959,5007, [N II] 6583, [S II] 6716,6731 + [O II] 7319,7330 + [O I] 6300,6364, and [S III] 9069,9532. About 3/4th of the remnant's main shell was imaged in all four filters. Considerable detail is observed in the reverse-shocked ejecta with typical knot scale lengths of 0.2"-0.4" (1 - 2 x 10^16 cm). Both bright and faint emission features appear highly clumped. Large differences in [S III] and [O III] line intensities indicating chemical abundance differences are also seen, particularly in knots located along the bright northern limb and near the base of the northeast jet. A line of curved overlapping filament in the remnant's northwestern rim appears to mark the location of the remnant's reverse shock front in this region. Finger-like ejecta structures elsewhere suggest cases where the reverse shock front is encountering the remnant's clumped ejecta. Narrow-band [N II] images of the remnant's circumstellar knots ("QSFs") reveal them to be 0.1"-0.6" thick knots and filaments, often with diffuse edges facing away from the center of expansion. Three color composite images of the whole remnant and certain sections along with individual filter enlargements of selected regions of the bright optical shell are presented and discussed.Comment: 26 pages, 12 figures Accepted to the Astronomical Journa