title: GoldenBraid: an iterative cloning system for standardized assembly of reusable genetic modules.
abstract: Synthetic Biology requires efficient and versatile DNA assembly systems to facilitate the building of new genetic modules/pathways from basic DNA parts in a standardized way. Here we present GoldenBraid (GB), a standardized assembly system based on type IIS restriction enzymes that allows the indefinite growth of reusable gene modules made of standardized DNA pieces. The GB system consists of a set of four destination plasmids (pDGBs) designed to incorporate multipartite assemblies made of standard DNA parts and to combine them binarily to build increasingly complex multigene constructs. The relative position of type IIS restriction sites inside pDGB vectors introduces a double loop ("braid") topology in the cloning strategy that allows the indefinite growth of composite parts through the succession of iterative assembling steps, while the overall simplicity of the system is maintained. We propose the use of GoldenBraid as an assembly standard for Plant Synthetic Biology. For this purpose we have GB-adapted a set of binary plasmids for A. tumefaciens-mediated plant transformation. Fast GB-engineering of several multigene T-DNAs, including two alternative modules made of five reusable devices each, and comprising a total of 19 basic parts are also described.
authors.affiliation: Instituto de Biología Molecular y Celular de Plantas, Consejo Superior de Investigaciones Científicas, Universidad Politécnica de Valencia, Valencia, Spain.
authors.forename: Alejandro
authors.initials: A
authors.lastname: Sarrion-Perdigones
authors.forename: Erica Elvira
authors.initials: EE
authors.lastname: Falconi
authors.forename: Sara I
authors.initials: SI
authors.lastname: Zandalinas
authors.forename: Paloma
authors.initials: P
authors.lastname: Juárez
authors.forename: Asun
authors.initials: A
authors.lastname: Fernández-del-Carmen
authors.forename: Antonio
authors.initials: A
authors.lastname: Granell
authors.forename: Diego
authors.initials: D
authors.lastname: Orzaez
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods*; D004247:DNA / Q000235:genetics* / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D007763:Lac Operon / Q000235:genetics; D008957:Models, Genetic; D030821:Plants, Genetically Modified; D010957:Plasmids / Q000235:genetics*; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D014026:Nicotiana / Q000235:genetics; D014170:Transformation, Genetic
chemical_list: D004247:DNA; D015252:Deoxyribonucleases, Type II Site-Specific