An Optimized Genotyping Workflow for Identifying Highly SCRaMbLEd Synthetic Yeasts.

Synthetic Sc2.0 yeast strains contain hundreds to thousands of recombination sites that allow restructuring of the genome by SCRaMbLE. Thus, a highly diverse yeast population can arise from a single genotype.

PhiReX 2.0: A Programmable and Red Light-Regulated CRISPR-dCas9 System for the Activation of Endogenous Genes in .

Metabolic engineering approaches do not exclusively require fine-tuning of heterologous genes but oftentimes also modulation or even induction of host gene expression, , in order to rewire metabolic fluxes. Here, we introduce the programmable red light switch PhiReX 2.0, which can rewire metabolic fluxes by targeting endogenous promoter sequences through single-guide RNAs (sgRNAs) and activate gene expression in upon red light stimulation.

The AssemblX Toolkit for Reliable and User-Friendly Multigene Assemblies.

The implementation of complex cloning projects covering the assembly of entire biological pathways or large genetic circuits poses a major challenge in the field of biotechnology and synthetic biology, as such projects can be costly and time-consuming. To overcome these difficulties, we developed the software-assisted AssemblX toolkit, which allows even unexperienced users to design, build, and subsequently test large DNA constructs. Currently, AssemblX allows the assembly of up to 25 functional units (e.

L-SCRaMbLE as a tool for light-controlled Cre-mediated recombination in yeast.

The synthetic yeast genome constructed by the International Synthetic Yeast Sc2.0 consortium adds thousands of loxPsym recombination sites to all 16 redesigned chromosomes, allowing the shuffling of Sc2.0 chromosome parts by the Cre-loxP recombination system thereby enabling genome evolution experiments.

PhiReX: a programmable and red light-regulated protein expression switch for yeast.

Highly regulated induction systems enabling dose-dependent and reversible fine-tuning of protein expression output are beneficial for engineering complex biosynthetic pathways. To address this, we developed PhiReX, a novel red/far-red light-regulated protein expression system for use in Saccharomyces cerevisiae. PhiReX is based on the combination of a customizable synTALE DNA-binding domain, the VP64 activation domain and the light-sensitive dimerization of the photoreceptor PhyB and its interacting partner PIF3 from Arabidopsis thaliana.

AssemblX: a user-friendly toolkit for rapid and reliable multi-gene assemblies.

The assembly of large DNA constructs coding for entire pathways poses a major challenge in the field of synthetic biology. Here, we present AssemblX, a novel, user-friendly and highly efficient multi-gene assembly strategy. The software-assisted AssemblX process allows even unexperienced users to rapidly design, build and test DNA constructs with currently up to 25 functional units, from 75 or more subunits.

Characterizing seamless ligation cloning extract for synthetic biological applications.

Synthetic biology aims at designing and engineering organisms. The engineering process typically requires the establishment of suitable DNA constructs generated through fusion of multiple protein coding and regulatory sequences. Conventional cloning techniques, including those involving restriction enzymes and ligases, are often of limited scope, in particular when many DNA fragments must be joined or scar-free fusions are mandatory.