An in vitro transposon system for highly regulated gene expression: construction of Escherichia coli strains with arabinose-dependent growth at low temperatures.

Placing a gene of interest under the control of an inducible promoter greatly aids the purification, localization and functional analysis of proteins but usually requires the sub-cloning of the gene of interest into an appropriate expression vector. Here, we describe an alternative approach employing in vitro transposition of Tn Omega P(BAD) to place the highly regulable, arabinose inducible P(BAD) promoter upstream of the gene to be expressed. The method is rapid, simple and facilitates the optimization of expression by producing constructs with variable distances between the P(BAD) promoter and the gene. To illustrate the use of this approach, we describe the construction of a strain of Escherichia coli in which growth at low temperatures on solid media is dependent on threshold levels of arabinose. Other uses of the transposable promoter are also discussed.

Quasi-static three-point bending of carbon fiber sandwich beams with square honeycomb cores

Sandwich beams comprising identical face sheets and a square honeycomb core were manufactured from carbon fiber composite sheets. Analytical expressions were derived for four competing collapse mechanisms of simply supported and clamped sandwich beams in three-point bending: core shear, face microbuckling, face wrinkling, and indentation. Selected geometries of sandwich beams were tested to illustrate these collapse modes, with good agreement between analytic predictions and measurements of the failure load. Finite element (FE) simulations of the three-point bending responses of these beams were also conducted by constructing a FE model by laying up unidirectional plies in appropriate orientations. The initiation and growth of damage in the laminates were included in the FE calculations. With this embellishment, the FE model was able to predict the measured load versus displacement response and the failure sequence in each of the composite beams. © 2011 American Society of Mechanical Engineers.