Biotechnology Letters

, Volume 31, Issue 10, pp 1577–1581 | Cite as

Construction of intragenic synthetic riboswitches for detection of a small molecule

Original Research Paper

Abstract

Bacterial sensors, based on ligand-mediated genetic control systems, are promising for on-site chemical detection because sensing targets and generating signals do not require costly instrumentation. Here, we have constructed intragenic synthetic riboswitches without relying on high-throughput screening and demonstrated that the riboswitches can be harnessed to develop bacterial sensors displaying readily visible reporter signals in response to theophylline. In vivo imaging using the riboswitch showed target-specific changes in the expression of a green fluorescence protein reporter, which was visible even to the naked eye.

Keywords

Aptamer Biosensor Riboswitch Synthetic biology Theophylline 

Notes

Acknowledgements

Authors are grateful to Ms. S.-H. Ryu for the technical assistance to preliminary study. This work was supported by BK21 program from the Korean Ministry of Education, start-up research funds provided by Yonsei University and Seoul R&BD Program (NT080612, KU080657).

References

  1. Breaker RR (2004) Natural and engineered nucleic acids as tools to explore biology. Nature 432:838–845PubMedCrossRefGoogle Scholar
  2. Bunka DHJ, Stockley PG (2006) Aptamers come of age at last. Nature Rev Microbiol 4:588–596CrossRefGoogle Scholar
  3. Cheah MT, Wachter A, Sudarsan N, Breaker RR (2007) Control of alternative RNA splicing and gene expression by eukaryotic riboswitches. Nature 447:497–500PubMedCrossRefGoogle Scholar
  4. Davidson EA, Ellington AD (2005) Engineering regulatory RNAs. Trends Biotechnol 23:109–112PubMedCrossRefGoogle Scholar
  5. Guido NJ, Wang X, Adalsteinsson D, McMillen D, Hasty J, Cantor CR, Elston TC, Collins JJ (2006) A bottom-up approach to gene regulation. Nature 439:856–860PubMedCrossRefGoogle Scholar
  6. Isaacs FJ, Dwyer DJ, Collins JJ (2006) RNA synthetic biology. Nature Biotechnol 24:545–554CrossRefGoogle Scholar
  7. Jenison RD, Gill SC, Pardi A, Polsky B (1994) High-resolution molecular discrimination by RNA. Science 263:1425–1429PubMedCrossRefGoogle Scholar
  8. Kozak M (2005) Regulation of translation via mRNA structure in prokaryotes and eukaryotes. Gene 361:13–37PubMedCrossRefGoogle Scholar
  9. Lutz R, Bujard H (1997) Independent and tight regulation of transcriptional units in Escherichia coli via the LacR/O, the TetR/O and AraC/I1-I2 regulatory elements. Nucleic Acids Res 25:1203–1210PubMedCrossRefGoogle Scholar
  10. Morita MT, Tanaka Y, Kodama TS, Kyogoku Y, Yanagi H, Yura T (1999) Translational induction of heat shock transcription factor σ32: evidence for a built-in RNA thermosensor. Genes Dev 13:655–665PubMedCrossRefGoogle Scholar
  11. Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory Press, Cold Spring HarborGoogle Scholar
  12. Sprinzak D, Elowitz MB (2005) Reconstruction of genetic circuits. Nature 438:443–448PubMedCrossRefGoogle Scholar
  13. Yusupova GZ, Yusupov MM, Cate JHD, Nodler HF (2001) The path of messenger RNA through the ribosome. Cell 106:233–241PubMedCrossRefGoogle Scholar
  14. Zimmermann GR, Wick CL, Shields TP, Jenison RD, Pardi A (2000) Molecular interactions and metal binding in the theophylline-binding core of an RNA aptamer. RNA 6:659–667PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2009

Authors and Affiliations

  1. 1.Department of BiotechnologyYonsei UniversitySeoulSouth Korea

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