Design and application of stationary phase combinatorial promoters

Current bacterial synthetic circuits rely on the fast dilution and high protein expression that occurs during exponential phase. However, constant exponential phase is both difficult to ensure in a lab environment and almost certainly impractical in any natural setting. Here, we characterize the performance of 13 E. coli native σ38 promoters, as well as a previously identified σ38 consensus promoter. We then make tetO combinatorial versions of the three strongest promoters to allow for inducible delayed expression. The design of these combinatorial promoters allows for design of circuits with inducible stationary phase activity that can be used for phase-dependent delays in dynamic circuits or spatial partitioning of biofilms

Clinical Legal Education in Asia - Accessing Justice for the Underprivileged

edited by Shuvro Sarker (Palgrave Macmillan), 2015, 288pp, £65 (hardback), ISBN 978-1-137-51752-

Three Poems

The doctor asks where it hurts. With the tip of a borrowed pen you trace the absent rib, that Biblical scar, tickle the edge of the raised ridge, a vacant dune between land and sea just below your left breast— like asking where pleasure starts, where whiskey settles, when you first knew his hand in your heart.https://digitalcommons.butler.edu/onearth/1032/thumbnail.jp

Assessment– Are Grade Descriptors the Way Forward?

The debate on how best to assess clinic, or indeed if it should be assessed at all has raged for decades and shows no sign of abating. The passage of time has been unable to resolve the question of assessment, no doubt due in part to the expansion and diversification of clinical legal education. The scope of clinic and its role in both society and as a teaching method is constantly evolving and assessment methods must develop to reflect the ever changing clinical profile. In an attempt to bring its assessment regime up to date, in 2007/2008 Northumbria University’s Student Law Office modified its assessment regime, replacing outmoded criteria with grade descriptors. This paper focuses on the use of grade descriptors and criterion referenced assessment in clinical legal education, addressing whether clinic should be assessed and which of the two methods is best suited to clinical legal education. The article draws on the experiences of clinicians and students to determine what issues this change in assessment regime has raised for the assessors and the student body. It concludes that it is appropriate to assess clinic by fully grading and suggests grade descriptors are the way forward

Quantitative Modeling of Integrase Dynamics Using a Novel Python Toolbox for Parameter Inference in Synthetic Biology

The recent abundance of high-throughput data for biological circuits enables data-driven quantitative modeling and parameter estimation. Common modeling issues include long computational times during parameter estimation, and the need for many iterations of this cycle to match data. Here, we present BioSCRAPE (Bio-circuit Stochastic Single-cell Reaction Analysis and Parameter Estimation) - a Python package for fast and flexible modeling and simulation for biological circuits. The BioSCRAPE package can be used for deterministic or stochastic simulations and can incorporate delayed reactions, cell growth, and cell division. Simulation run times obtained with the package are comparable to those obtained using C code - this is particularly advantageous for computationally expensive applications such as Bayesian inference or simulation of cell lineages. We first show the package's simulation capabilities on a variety of example simulations of stochastic gene expression. We then further demonstrate the package by using it to do parameter inference for a model of integrase dynamics using experimental data. The BioSCRAPE package is publicly available online along with more detailed documentation and examples

Control Theory for Synthetic Biology: Recent Advances in System Characterization, Control Design, and Controller Implementation for Synthetic Biology

Living organisms are differentiated by their genetic material-millions to billions of DNA bases encoding thousands of genes. These genes are translated into a vast array of proteins, many of which have functions that are still unknown. Previously, it was believed that simply knowing the genetic sequence of an organism would be the key to unlocking all understanding. However, as DNA sequencing technology has become affordable, it has become clear that living cells are governed by complex, multilayered networks of gene regulation that cannot be deduced from sequence alone. Synthetic biology as a field might best be characterized as a learn-by-building approach, in which scientists attempt to engineer molecular pathways that do not exist in nature. In doing so, they test the limits of both natural and engineered organisms

Design and application of stationary phase combinatorial promoters

Current bacterial synthetic circuits rely on the fast dilution and high protein expression that occurs during exponential phase. However, constant exponential phase is both difficult to ensure in a lab environment and almost certainly impractical in any natural setting. Here, we characterize the performance of 13 E. coli native σ38 promoters, as well as a previously identified σ38 consensus promoter. We then make tetO combinatorial versions of the three strongest promoters to allow for inducible delayed expression. The design of these combinatorial promoters allows for design of circuits with inducible stationary phase activity that can be used for phase-dependent delays in dynamic circuits or spatial partitioning of biofilms

Engineering pulsatile communication in bacterial consortia

Lux-type quorum sensing systems enable communication in bacteria with only two protein components: a signal synthase and an inducible transcription activator. The simplicity of these systems makes them a popular choice for engineering collaborative behaviors in synthetic bacterial consortia, such as photographic edge detection and synchronized oscillation. To add to this body of work, we propose a pulsatile communication circuit that enables dynamic patterning and long-distance communication analogous to action potentials traveling through nerve tissue. We employed a model-driven design paradigm involving incremental characterization of in vivo design candidates with increasing circuit complexity. Beginning with a simple inducible reporter system, we screened a small number of circuits varying in their promoter and ribosomal binding site strengths. From this candidate pool, we selected a candidate to be the seed network for the subsequent round of more complex circuit variants, likewise variable in promoter and RBS strengths. The selection criteria at each level of complexity is tailored to optimize a different desirable performance characteristic. By this approach we individually optimized reporter signal-to-background ratio, pulsatile response to induction, and quiescent basal transcription, avoiding large library screens while ensuring robust performance of the composite circuit

Ive Got My Virtual Eye On You: Remote Proctors And Academic Integrity

This paper discusses the challenges of online teaching, the reasons students cheat and one means of curtailing that cheating in the online environment. The use of Securexam Remote Proctor System in one university application is reviewed

Fast and flexible simulation and parameter estimation for synthetic biology using bioscrape

In systems and synthetic biology, it is common to build chemical reaction network (CRN) models of biochemical circuits and networks. Although automation and other high-throughput techniques have led to an abundance of data enabling data-driven quantitative modeling and parameter estimation, the intense amount of simulation needed for these methods still frequently results in a computational bottleneck. Here we present bioscrape (Bio-circuit Stochastic Single-cell Reaction Analysis and Parameter Estimation) - a Python package for fast and flexible modeling and simulation of highly customizable chemical reaction networks. Specifically, bioscrape supports deterministic and stochastic simulations, which can incorporate delay, cell growth, and cell division. All functionalities - reaction models, simulation algorithms, cell growth models, and partioning models - are implemented as interfaces in an easily extensible and modular object-oriented framework. Models can be constructed via Systems Biology Markup Language (SBML), a simple internal XML language, or specified programmatically via a Python API. Simulation run times obtained with the package are comparable to those obtained using C code - this is particularly advantageous for computationally expensive applications such as Bayesian inference or simulation of cell lineages. We first show the package's simulation capabilities on a variety of example simulations of stochastic gene expression. We then further demonstrate the package by using it to do parameter inference on a model of integrase enzyme-mediated DNA recombination dynamics with experimental data. The bioscrape package is publicly available online (https://github.com/ananswam/bioscrape) along with more detailed documentation and examples