ABSTRACT
Synthetic biology aims to engineer and redesign biological systems for useful real-world applications in biomanufacturing, biosensing and biotherapy following a typical design-build-test cycle. Inspired from computer science and electronics, synthetic gene circuits have been designed to exhibit control over the flow of information in biological systems. Two types are Boolean logic inspired TRUE or FALSE digital logic and graded analog computation. Key principles for gene circuit engineering include modularity, orthogonality, predictability and reliability. Initial circuits in the field were small and hampered by a lack of modular and orthogonal components, however in recent years the library of available parts has increased vastly. New tools for high throughput DNA assembly and characterization have been developed enabling rapid prototyping, systematic in situ characterization, as well as automated design and assembly of circuits. Recently implemented computing paradigms in circuit memory and distributed computing using cell consortia will also be discussed. Finally, we will examine existing challenges in building predictable large-scale circuits including modularity, context dependency and metabolic burden as well as tools and methods used to resolve them. These new trends and techniques have the potential to accelerate design of larger gene circuits and result in an increase in our basic understanding of circuit and host behaviour.
ABSTRACT
Efficiency of yeast transformation is determined by the rate of yeast endocytosis. The aim of this study was to investigate the effect of introducing amino acids and other nutrients (inositol, adenine, or p-aminobenzoic acid) in the transformation medium to develop a highly efficient yeast transformation protocol. The target of rapamycin complex 1 (TORC1) kinase signalling complex influences the rate of yeast endocytosis. TORC signaling is induced by amino acids in the media. Here, we found that increasing the concentration of amino acids and other nutrients in the growth media lead to an increase yeast transformation efficiency up to 107 CFU per µg plasmid DNA and per 108 cells with a 13.8 kb plasmid DNA. This is over 130 times that of current published methods. This improvement may facilitate more efficient experimentation in which transformation efficiency is critical, such as yeast two-hybrid screening.
