Collaborative Research: Synthetic and Systems Biology Approaches to Semi-synthetic Cells with Expanded DNA Alphabets

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This project focuses on novel forms of DNA that are not built only from the standard four nucleotides found in natural DNA, but with two or four additional nucleotides, to form an "Artificially Expanded Genetic Information System" (AEGIS). Expanded DNA has already seen widespread use in human diagnostics and environmental surveillance for pathogens. This project will break new ground in developing our understanding of DNA, and develop new technology for business, defense, and health. It will do so by studying the interaction of AEGIS DNA and its components with living microbial cells. As AEGIS can support Darwinian evolution, seen by many as the essence of life, this project will drive a deeper understanding of life. Further, it will allow laboratory evolution of new functional materials, adding evolution and adaptation to points in biotechnology where description and design are inadequate. Finally, this project emphasizes public outreach efforts and the training of early career scientists. This project will complete engineering of bacterial strains that replicate, evolve, and use DNA built not from 4 different nucleotides, but from 6 or 8, DNA in an Artificially Expanded Genetic Information System (AEGIS). These strains, "Second Examples of Genetics Undergoing Evolution" (SEGUE) will use one (or both) of two 6-letter genetic alphabets, GACTKX and GACTZP. A multidisciplinary team of chemists, molecular biologists, philosophers, and bioengineers will exploit comprehensive preliminary data. Here, critical points are integrated to yield results beneficial across multiple platforms:• Broader Impacts in training, outreach, and risk analysis by having direct contact between experiments and those who study the history of and "science of" science.• Work to develop polymerases that replicate AEGIS DNA in vitro and in vivo SEGUE. These will themselves have value in environmental pathogen surveillance, diagnostics, and therapeutic development, as well as the creation of new materials for biotechnology.• Protein engineering to build a metabolic pathway to biosynthesize, inside of living cells, AEGIS triphosphates, allowing AEGIS to be used inexpensively.• Study of interactions between AEGIS and living cells, specifically to identify "third party" genes and regulatory systems that improve the performance and lower the cost of AEGIS in SEGUE.• Broader impacts include the training of experimental scientists in philosophical and ethical features of synthetic biology, iGEM projects, and training for undergraduate and graduate students. The implications throughout science are enormous, and touch on concepts of shadow biospheres, life on Mars and elsewhere in the Solar System, and how life in general is conceived and understood by scientists and the general public alike. Two updated books will provide a framework for thinking about these and other issues, Cleland's "The Quest for a Universal Theory of Life" and Benner's "Life, the Universe, and the Scientific Method." Other outreach materials will include the blog: "Primordial Scoop", which discusses science and technology from philosophical and ethical perspectives. Educational materials will be internationally available, including in classrooms, museums, and planetariums.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.