Skip to main content

Engineering the Logical Properties of a Genetic AND Gate

  • Protocol
  • First Online:
Nanoscale Biocatalysis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 743))

Abstract

Synthetic biology promises to enhance our ability to control biological systems by creating a systematic approach for the construction of genetic circuits that reliably program cellular function. As part of this approach, efficient methods are needed for the tuning of genetic circuits so as to allow for optimization of a design despite varying cellular contexts and incomplete understanding of in vivo biological interactions. Here we outline an optimization method that we have used to improve the logical responses of a genetic AND logic gate derived from components of the LuxI–LuxR bacterial quorum-sensing system. Basing our approach on the idea of evolutionary design, we improved the properties of our genetic AND logic gate by using directed evolution and a two-step screening process to alter the activities of the LuxR transcriptional activator. Using this method, we were able to rapidly enhance the AND gate’s logical responses and have increased the specificities of these responses by ∼1.5-fold.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Andrianantoandro, E., Basu, S., Karig, D. K., and Weiss, R. (2006) Synthetic biology: New engineering rules for an emerging discipline. Mol. Syst. Biol. 2, 1–14.

    Article  Google Scholar 

  2. Marguet, P., Balagadde, F., Tan, C., and You, L. (2007) Biology by design: Reduction and synthesis of cellular components and behaviour. J. R. Soc. Interface 4, 607–623.

    Article  CAS  Google Scholar 

  3. Haseltine, E. L., and Arnold, F. H. (2007) Synthetic gene circuits: Design with directed evolution. Annu. Rev. Biophys. Biomol. Struct. 36, 1–19.

    Article  CAS  Google Scholar 

  4. Yokobayashi, Y., Weiss, R., and Arnold, F. H. (2002) Directed evolution of a genetic circuit. Proc. Natl. Acad. Sci. USA 99, 16587–16591.

    Article  CAS  Google Scholar 

  5. Chockalingam, K., Chen, Z. L., Katzenellenbogen, J. A., and Zhao, H. M. (2005) Directed evolution of specific receptor-ligand pairs for use in the creation of gene switches. Proc. Natl. Acad. Sci. USA 102, 5691–5696.

    Article  CAS  Google Scholar 

  6. Alper, H., Fischer, C., Nevoigt, E., and Stephanopoulos, G. (2005) Tuning genetic control through promoter engineering. Proc. Natl. Acad. Sci. USA 102, 12678–12683.

    Article  CAS  Google Scholar 

  7. Whitehead, N. A., Barnard, A. M. L., Slater, H., Simpson, N. J. L., and Salmond, G. P. C. (2001) Quorum-sensing in gram-negative bacteria. FEMS Microbiol. Rev. 25, 365–404.

    Article  CAS  Google Scholar 

  8. Urbanowski, A. L., Lostroh, C. P., and Greenberg, E. P. (2004) Reversible acyl-homoserine lactone binding to purified Vibrio fischeri LuxR protein. J. Bacteriol. 186, 631–637.

    Article  CAS  Google Scholar 

  9. Gilbert, W., and Muller-Hill, B. (1970) The lactose repressor. In The Lactose Operon (Beckwith, J. and Zipser, D., Eds.), Cold Spring Harbor Lab Press, Plainview, NY, pp. 93–109.

    Google Scholar 

  10. Sayut, D. J., Niu, Y., and Sun, L. (2009) Construction and enhancement of a minimal genetic AND logic gate. Appl. Environ. Microbiol. 75, 637–642.

    Article  CAS  Google Scholar 

  11. Cadwell, R. C., and Joyce, G. F. (1994) Mutagenic PCR. PCR Methods Appl. 3, S136–S140.

    CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by Grant # CBET-0747728 from the National Science Foundation and Grant # IRG 93-033 from the American Cancer Society.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Lianhong Sun .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2011 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Sayut, D.J., Niu, Y., Sun, L. (2011). Engineering the Logical Properties of a Genetic AND Gate. In: Wang, P. (eds) Nanoscale Biocatalysis. Methods in Molecular Biology, vol 743. Humana Press. https://doi.org/10.1007/978-1-61779-132-1_14

Download citation

  • DOI: https://doi.org/10.1007/978-1-61779-132-1_14

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-131-4

  • Online ISBN: 978-1-61779-132-1

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics