title: The spinach RNA aptamer as a characterization tool for synthetic biology.
abstract: Characterization of genetic control elements is essential for the predictable engineering of synthetic biology systems. The current standard for in vivo characterization of control elements is through the use of fluorescent reporter proteins such as green fluorescent protein (GFP). Gene expression, however, involves not only protein production but also the production of mRNA. Here, we present the use of the Spinach aptamer sequence, an RNA mimic of GFP, as a tool to characterize mRNA expression in Escherichia coli. We show how the aptamer can be incorporated into gene expression cassettes and how co-expressing it with a red fluorescent protein (mRFP1) allows, for the first time, simultaneous measurement of mRNA and protein levels from engineered constructs. Using flow cytometry, we apply this tool here to evaluate ribosome binding site sequences and promoters and use it to highlight the differences in the temporal behavior of transcription and translation.
authors.affiliation: Centre for Synthetic Biology and Innovation, Imperial College London , South Kensington Campus, Exhibition Rd, London SW7 2AZ, United Kingdom.
authors.forename: Georgios
authors.initials: G
authors.lastname: Pothoulakis
authors.forename: Francesca
authors.initials: F
authors.lastname: Ceroni
authors.forename: Benjamin
authors.initials: B
authors.lastname: Reeve
authors.forename: Tom
authors.initials: T
authors.lastname: Ellis
mesh_terms: D052157:Aptamers, Nucleotide / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005818:Genetic Engineering / Q000379:methods*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D012313:RNA / Q000235:genetics* / Q000378:metabolism; D012329:RNA, Bacterial / Q000235:genetics / Q000378:metabolism; D018724:Spinacia oleracea / Q000235:genetics*; D058615:Synthetic Biology / Q000379:methods*
chemical_list: D052157:Aptamers, Nucleotide; D008164:Luminescent Proteins; D012329:RNA, Bacterial; D012313:RNA