von Dassow, P. | bioRxiv Science

bioRxiv2019

Universal Loop assembly (uLoop): open, efficient, and species-agnostic DNA fabrication

Standardised Type IIS DNA assembly methods are becoming essential for biological engineering and research. Although a common syntax has been proposed to enable higher interoperability between DNA libraries, Golden Gate (GG)-based assembly systems remain specific to target organisms. Furthermore, these GG assembly systems become laborious and unnecessarily complicated beyond the assembly of 4 transcriptional units. Here, we describe \"universal Loop\" (uLoop) assembly, a simple system based on Loop assembly that enables hierarchical fabrication of large DNA constructs (> 30 kb) for any organism of choice. uLoop comprises two sets of four plasmids that are iteratively used as odd and even levels to compile DNA elements in an exponential manner (4n-1). The elements required for transformation/maintenance in target organisms are also assembled as standardised parts, enabling customisation of host-specific plasmids. Thus, this species-agnostic method decouples efficiency of assembly from the stability of vectors in the target organism. As a proof-of-concept, we show the engineering of multi-gene expression vectors in diatoms, yeast, plants and bacteria. These resources will become available through the OpenMTA for unrestricted sharing and open-access.\n\n\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=144 SRC=\"FIGDIR/small/744854v2_ufig.gif\" ALT=\"Figure 1\">\nView larger version (31K):\norg.highwire.dtl.DTLVardef@c36f4forg.highwire.dtl.DTLVardef@c0b007org.highwire.dtl.DTLVardef@64c810org.highwire.dtl.DTLVardef@8a94b0_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2018

Strength in numbers: collaborative science for new experimental model systems

Our current understanding of biology is heavily based on the contributions from a small number of genetically tractable model organisms. Most eukaryotic phyla lack such experimental models, and this limits our ability to explore the molecular mechanisms that ultimately define their biology, ecology, and diversity. In particular, marine protists suffer from a paucity of model organisms despite playing critical roles in global nutrient cycles, food webs, and climate. To address this deficit, an initiative was launched in 2015 to foster development of ecologically and taxonomically diverse marine protist genetic models. This multifaceted, complex but important challenge required a highly collaborative community-based approach. Herein we describe this approach, the advances achieved, and the lessons learned by participants in this novel community-based model for research.

microbiology↗