title: DNA-functionalized hydrogels for confined membrane-free in vitro transcription/translation.
abstract: We microfluidically fabricate bio-orthogonal DNA-functionalized porous hydrogels from hyaluronic acid that are employed in in vitro transcription/translation (IVTT) of a green fluorescent protein. By co-encapsulating individual hydrogel particles and the IVTT machinery in water-in-oil microdroplets, we study protein expression in a defined reaction volume. Our approach enables precise control over protein expression rates by gene dosage. We show that gene transcription and translation are confined to the membrane-free hydrogel matrix, which contributes to the design of membrane-free protocells.
authors.affiliation: Radboud University Nijmegen, Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands. j.thiele@science.ru.nl.
authors.forename: J
authors.initials: J
authors.lastname: Thiele
authors.forename: Y
authors.initials: Y
authors.lastname: Ma
authors.forename: D
authors.initials: D
authors.lastname: Foschepoth
authors.forename: M M K
authors.initials: MM
authors.lastname: Hansen
authors.forename: C
authors.initials: C
authors.lastname: Steffen
authors.forename: H A
authors.initials: HA
authors.lastname: Heus
authors.forename: W T S
authors.initials: WT
authors.lastname: Huck
mesh_terms: D002474:Cell-Free System; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D004129:Dimethylpolysiloxanes / Q000737:chemistry; D004655:Emulsions; D004867:Equipment Design; D029968:Escherichia coli Proteins / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006820:Hyaluronic Acid / Q000737:chemistry; D020100:Hydrogels / Q000737:chemistry*; D061925:Immobilized Nucleic Acids / Q000737:chemistry* / Q000378:metabolism; D008422:Materials Testing; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D008863:Microspheres; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016062:Porosity; D066330:Printing, Three-Dimensional; D014176:Protein Biosynthesis; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013499:Surface Properties; D014158:Transcription, Genetic
chemical_list: D004129:Dimethylpolysiloxanes; D004655:Emulsions; D029968:Escherichia coli Proteins; D020100:Hydrogels; D061925:Immobilized Nucleic Acids; D010446:Peptide Fragments; D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; C013830:baysilon; D006820:Hyaluronic Acid; D004247:DNA