In 2010 the J. Craig Venter Institute (JCVI) shocked the scientific world by creating the first synthetic cell, Mycoplasma mycoides JCVI-syn1.0, which contained slightly over 1 million base pairs and 901 genes. This was a significant achievement in the field of synthetic biology and allowed scientists to ask questions about which genes and metabolic pathways were absolutely essential for sustaining organismal life processes. Yet, the researchers felt they could improve upon their original design and strip away even more genes without compromising the cell's overall health.
Now, the team of scientists from JCVI that were involved with version 1.0 teamed up with researchers from Synthetic Genomics (SGI) to assemble the first minimal synthetic bacterial cell, which they dubbed JCVI-syn3.0. Using JCVI-syn1.0 as a template, the investigators created an organism that has almost half the number of base pairs (531,560) of the original and a total of 473 genes, making it the smallest genome of any organism that can be grown in laboratory media. Astonishingly, of these genes, 149 are of unknown biological function.
“Our attempt to design and create a new species, while ultimately successful, revealed that 32% of the genes essential for life in this cell are of unknown function, and showed that many are highly conserved in numerous species. All the bioinformatics studies over the past 20 years have underestimated the number of essential genes by focusing only on the known world. This is an important observation that we are carrying forward into the study of the human genome,” explained senior study author Craig Venter, Ph.D., founder, executive chairman, and CEO of JCVI.
To create JCVI-syn3.0, the team used an approach of whole genome design and chemical synthesis, followed by genome transplantation to test if the cell was viable. Their first attempt to minimize the genome began with a simple approach using information in the biochemical literature and some limited transposon mutagenesis work, but this did not result in a viable genome. After improving the transposon methods, the researchers discovered a set of quasi-essential genes that are necessary for robust growth—explaining the failure of their initial attempt.
The JCVI/SGI team built the genome in eight overlapping segments so that each could be tested separately before combining them to generate a minimal genome. Moreover, the researchers went through three cycles of designing, building, and testing to ensure that the quasi-essential genes remained, which in the end resulted in a viable, self-replicating minimal synthetic cell that contained just 473 genes, 35 of which are RNA coding. In addition, the cell contains a unique 16S gene sequence.
The findings from this study were published recently in Science in an article entitled “Design and Synthesis of a Minimal Bacterial Genome.”
Dr. Venter and his colleagues were able to assign biological function to the majority of the genes, with 41% of them responsible for genome expression information, 18% related to cell membrane structure and function, 17% related to cytosolic metabolism, and 7% preservation of genome information. However, a surprising 149 genes could not be assigned a particular biological function despite intensive study—an area of continued work for the researchers.
“This paper represents more than five years of work by an amazingly talented group of people,” noted lead study author Clyde Hutchison, Ph.D., distinguished professor at JCVI. “Our goal is to have a cell for which the precise biological function of every gene is known.”
The researchers are hopeful that a primary outcome of the minimal cell program are new tools and semiautomated processes for whole genome synthesis, which would advance our understanding of life-sustaining metabolic processes.
“This paper signifies a major step toward our ability to design and build synthetic organisms from the bottom up with predictable outcomes. The tools and knowledge gained from this work will be essential to producing next generation production platforms for a broad range of disciplines,” concluded co-author Daniel Gibson, Ph.D., vice president of DNA Technologies at SGI and associate professor of synthetic biology at JCVI.