Methodology

Measuring the activity of BioBrick promoters using an in vivo reference standard

Jason R Kelly¹ , Adam J Rubin¹ , Joseph H Davis² , Caroline M Ajo-Franklin³ , John Cumbers⁴ , Michael J Czar⁵ , Kim de Mora⁶ , Aaron L Glieberman⁷ , Dileep D Monie^8,9 and Drew Endy^1,10

¹  Department of Biological Engineering, Massachusetts Institute of Technology, 77 Massachusetts Avenue, 68-580, Cambridge Massachusetts 02139, USA

²  Department of Biology, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge Massachusetts 02139, USA

³  Molecular Foundry, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA

⁴  Molecular Biology, Cell Biology, and Biochemistry, Brown University, Box G-W, Providence, Rhode Island, 02912, USA

⁵  Virginia Bioinformatics Institute, Virginia Polytechnic and State University, Washington Street, MC0477, Blacksburg, VA 24061, USA

⁶  Institute for Materials and Processes, The University of Edinburgh, King's Buildings, Mayfield Road, Edinburgh, Scotland EH9 3JL, UK

⁷  Biomedical Engineering, Division of Engineering, Brown University, Providence, Rhode Island 02912, USA

⁸  Department of Ophthalmology, Johns Hopkins University School of Medicine, 600 N Wolfe St, Baltimore, Maryland 21287, USA

⁹  Profectus Biosciences, Inc., 6411 Beckley St, Baltimore, Maryland 21224, USA

¹⁰  Department of Bioengineering, Stanford University, Stanford California 94305 USA

author email corresponding author email

Journal of Biological Engineering 2009, 3:4doi:10.1186/1754-1611-3-4

Published:	20 March 2009

Abstract

Background

The engineering of many-component, synthetic biological systems is being made easier by the development of collections of reusable, standard biological parts. However, the complexity of biology makes it difficult to predict the extent to which such efforts will succeed. As a first practical example, the Registry of Standard Biological Parts started at MIT now maintains and distributes thousands of BioBrick™ standard biological parts. However, BioBrick parts are only standardized in terms of how individual parts are physically assembled into multi-component systems, and most parts remain uncharacterized. Standardized tools, techniques, and units of measurement are needed to facilitate the characterization and reuse of parts by independent researchers across many laboratories.

Results

We found that the absolute activity of BioBrick promoters varies across experimental conditions and measurement instruments. We choose one promoter (BBa_J23101) to serve as an in vivo reference standard for promoter activity. We demonstrated that, by measuring the activity of promoters relative to BBa_J23101, we could reduce variation in reported promoter activity due to differences in test conditions and measurement instruments by ~50%. We defined a Relative Promoter Unit (RPU) in order to report promoter characterization data in compatible units and developed a measurement kit so that researchers might more easily adopt RPU as a standard unit for reporting promoter activity. We distributed a set of test promoters to multiple labs and found good agreement in the reported relative activities of promoters so measured. We also characterized the relative activities of a reference collection of BioBrick promoters in order to further support adoption of RPU-based measurement standards.

Conclusion

Relative activity measurements based on an in vivoreference standard enables improved measurement of promoter activity given variation in measurement conditions and instruments. These improvements are sufficient to begin to support the measurement of promoter activities across many laboratories. Additional in vivo reference standards for other types of biological functions would seem likely to have similar utility, and could thus improve research on the design, production, and reuse of standard biological parts.