Applied Cellular Physiology and Metabolic Engineering

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A novel counter-selection method for markerless genetic modification in Synechocystis sp. PCC 6803

  1. Yi Ern Cheah1,
  2. Stevan C. Albers2,
  3. Christie A. M. Peebles1,2,*

Article first published online: 4 DEC 2012

DOI: 10.1002/btpr.1661

Issue

Biotechnology Progress

Biotechnology Progress

Volume 29, Issue 1, pages 23–30, January/February 2013

Additional Information

How to Cite

Cheah, Y. E., Albers, S. C. and Peebles, C. A. M. (2013), A novel counter-selection method for markerless genetic modification in Synechocystis sp. PCC 6803. Biotechnol Progress, 29: 23–30. doi: 10.1002/btpr.1661

Author Information

  1. 1

    Dept. of Chemical and Biological Engineering, Colorado State University, Fort Collins, CO 80523

  2. 2

    Cell and Molecular Biology Graduate Program, Colorado State University, Fort Collins, CO 80523

*Department of Chemical and Biological Engineering, Colorado State University, Fort Collins, CO, USA 80523-1370

Publication History

  1. Issue published online: 4 FEB 2013
  2. Article first published online: 4 DEC 2012
  3. Accepted manuscript online: 1 NOV 2012 08:21AM EST
  4. Manuscript Revised: 15 OCT 2012
  5. Manuscript Received: 21 JUN 2012

Funded by

  • Sustainable Bioenergy Development Center at Colorado State University

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Keywords:

  • cyanobacteria;
  • Synechocystis;
  • homologous recombination;
  • markerless transformation;
  • genetic engineering;
  • nickel response system;
  • mazF

Abstract

The cyanobacterium Synechocystis sp. PCC 6803 is a photosynthetic organism capable of efficient harnessing of solar energy while capturing CO2 from the environment. Methods to genetically alter its genomic DNA are essential for elucidating gene functions and are useful tools for metabolic engineering. In this study, a novel counter-selection method for the genetic alteration of Synechocystis was developed. This method utilizes the nickel inducible expression of mazF, a general protein synthesis inhibitor, as a counter-selection marker. Counter-selection is particularly useful because the engineered strain is free of any markers which make further genetic modification independent of available antibiotic resistance genes. The usability of this method was further demonstrated by altering genes at several loci in two variants of Synechocystis. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2013

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