Broccoli aptamer allows quantitative transcription regulation studies in vitro

• Published in: PloS one Vol. 19; no. 6; p. e0304677
• Main Authors: van der Sijs, Amanda, Visser, Thomas, Moerman, Pepijn, Folkers, Gert, Kegel, Willem
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 13.06.2024
• Subjects: Aptamers; Aptamers, Nucleotide - chemistry; Aptamers, Nucleotide - metabolism; Binding sites; Biology and life sciences; Brassica - genetics; Brassica - metabolism; Broccoli; Delay time; E coli; Ethylenediaminetetraacetic acid; Gene expression; Gene Expression Regulation; Gene regulation; Genetic research; Genetic transcription; In vivo methods and tests; Lactose; Lactose repressor; Lagrange multiplier; Medicine and Health Sciences; Noncoding DNA; Physical Sciences; Proteins; Quantitative analysis; Quantitative research; Research and Analysis Methods; RNA polymerase; Thermodynamics; Transcription; Transcription factors; Transcription, Genetic; Vegetables
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0304677

Quantitative transcription regulation studies in vivo and in vitro often make use of reporter proteins. Here we show that using Broccoli aptamers, quantitative study of transcription in various regulatory scenarios is possible without a translational step. To explore the method we studied several regulatory scenarios that we analyzed using thermodynamic occupancy-based models, and found excellent agreement with previous studies. In the next step we show that non-coding DNA can have a dramatic effect on the level of transcription, similar to the influence of the lac repressor with a strong affinity to operator sites. Finally, we point out the limitations of the method in terms of delay times coupled to the folding of the aptamer. We conclude that the Broccoli aptamer is suitable for quantitative transcription measurements.