Toward a genetic system in the marine cyanobacterium Prochlorococcus

Raphaël Laurenceau; Christina Bliem; Marcia S Osburne; Jamie W Becker; Steven J Biller; Andres Cubillos-Ruiz; Sallie W Chisholm

doi:10.1099/acmi.0.000107

Toward a genetic system in the marine cyanobacterium Prochlorococcus

Access Microbiol. 2020 Feb 19;2(4):acmi000107. doi: 10.1099/acmi.0.000107. eCollection 2020.

Authors

Raphaël Laurenceau¹, Christina Bliem¹, Marcia S Osburne^{1

2}, Jamie W Becker^{1

3}, Steven J Biller^{1

4}, Andres Cubillos-Ruiz^{1

5

6

7}, Sallie W Chisholm^{1

8}

Affiliations

¹ Department of Civil and Environmental Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.
² Present address: Department of Molecular Biology and Microbiology Tufts University School of Medicine, Boston, MA, USA.
³ Present address: Department of Biology, Haverford College, Haverford, PA, USA.
⁴ Present address: Department of Biological Sciences, Wellesley College, Wellesley, MA, USA.
⁵ Present address: Institute for Medical Engineering and Science, Department of Biological Engineering, and Synthetic Biology Center, Massachusetts Institute of Technology, Cambridge, MA, USA.
⁶ Present address: Broad Institute of MIT and Harvard, Cambridge, MA, USA.
⁷ Present address: Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA, USA.
⁸ Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA.

Abstract

As the smallest and most abundant primary producer in the oceans, the cyanobacterium Prochlorococcus is of interest to diverse branches of science. For the past 30 years, research on this minimal phototroph has led to a growing understanding of biological organization across multiple scales, from the genome to the global ocean ecosystem. Progress in understanding drivers of its diversity and ecology, as well as molecular mechanisms underpinning its streamlined simplicity, has been hampered by the inability to manipulate these cells genetically. Multiple attempts have been made to develop an efficient genetic transformation method for Prochlorococcus over the years; all have been unsuccessful to date, despite some success with their close relative, Synechococcus . To avoid the pursuit of unproductive paths, we report here what has not worked in our hands, as well as our progress developing a method to screen the most efficient electroporation parameters for optimal DNA delivery into Prochlorococcus cells. We also report a novel protocol for obtaining axenic colonies and a new method for differentiating live and dead cells. The electroporation method can be used to optimize DNA delivery into any bacterium, making it a useful tool for advancing transformation systems in other genetically recalcitrant microorganisms.

Keywords: Prochlorococcus; genetic system; intractable bacteria.