Abstract
Photosynthetic organisms, and especially cyanobacteria, hold great promise as sources of renewably-produced fuels, bulk and specialty chemicals, and nutritional products. Synthetic biology tools can help unlock cyanobacteria's potential for these functions, but unfortunately tool development for these organisms has lagged behind that for S. cerevisiae and E. coli. While these organisms may in many cases be more difficult to work with as “chassis” strains for synthetic biology than certain heterotrophs, the unique advantages of autotrophs in biotechnology applications as well as the scientific importance of improved understanding of photosynthesis warrant the development of these systems into something akin to a “green E. coli.” In this review, we highlight unique challenges and opportunities for development of synthetic biology approaches in cyanobacteria. We review classical and recently developed methods for constructing targeted mutants in various cyanobacterial strains, and offer perspective on what genetic tools might most greatly expand the ability to engineer new functions in such strains. Similarly, we review what genetic parts are most needed for the development of cyanobacterial synthetic biology. Finally, we highlight recent methods to construct genome-scale models of cyanobacterial metabolism and to use those models to measure properties of autotrophic metabolism. Throughout this paper, we discuss some of the unique challenges of a diurnal, autotrophic lifestyle along with how the development of synthetic biology and biotechnology in cyanobacteria must fit within those constraints.
Highlights
Cyanobacteria have garnered a great deal of attention recently as biofuel-producing organisms
We will look toward how the techniques of the emerging field of synthetic biology might bear fruit in improving the output of such engineered strains
We believe that the tools of synthetic biology can help with this challenge
Summary
Cyanobacteria have garnered a great deal of attention recently as biofuel-producing organisms. As compared with eukaryotic algae and plants, cyanobacteria are much easier to manipulate genetically and grow much faster They have been engineered to produce a wide and ever-expanding range of products including fatty acids, long-chain alcohols, alkanes, ethylene, polyhydroxybutyrate, 2,3-Butanediol, ethanol, and hydrogen. Several methods have been developed that allow the creation of markerless mutations in cyanobacterial chromosomes (Figure 1B) Two of these methods operate on a similar principle: First, a conditionally toxic gene is linked to an antibiotic resistance cassette and inserted into the chromosome, GENETIC MODIFICATION IN CIS: CHROMOSOME EDITING. Cis genetic modification is the most common approach in cyanobacterial synthetic biology This approach takes advantage of the capability of many cyanobacterial strains for natural transformation and homologous recombination (see Table 1) to create insertion, deletion, or replacement mutations in cyanobacterial chromosomes. While such stability is desirable, systems that create major metabolic demand, by for example redirecting flux into biofuel-producing pathways, will face greater selective pressures for mutation or loss of heterologous genes
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