Researchers created over 4.3 billion combinatorial genomic variants (of E Coli) per day. They isolated variants with more than fivefold increase in lycopene production within 3 days, a significant improvement over existing metabolic engineering techniques.
Multiplex automated genome engineering (MAGE) is used for large-scale programming and evolution of cells. MAGE simultaneously targets many locations on the chromosome for modification in a single cell or across a population of cells, thus producing combinatorial genomic diversity. Because the process is cyclical and scalable, they constructed prototype devices that automate the MAGE technology to facilitate rapid and continuous generation of a diverse set of genetic changes (mismatches, insertions, deletions). We applied MAGE to optimize the 1-deoxy-d-xylulose-5-phosphate (DXP) biosynthesis pathway in Escherichia coli to overproduce the industrially important isoprenoid lycopene
The E. coli bacterium contains approximately 4,500 genes. The team focused on 24 of these—honing a pathway with tremendous potential—to increase production of the antioxidant, optimizing the sequences simultaneously. They took the 24 DNA sequences, divided them up into manageable 90-letter segments, and modified each, generating a suite of genetic variants. Next, armed with specific sequences, the team enlisted a company to manufacture thousands of unique constructs. The team was then able to insert these new genetic constructs back into the cells, allowing the natural cellular machinery to absorb this revised genetic material.
Some bacteria ended up with one construct, some ended up with multiple constructs. The resulting pool contained an assortment of cells, some better at producing lycopene than others. The team extracted the best producers from the pool and repeated the process over and over to further hone the manufacturing machinery. To make things easier, the researchers automated all of these steps.
“We accelerated evolution, generating as many as 15 billion genetic variants in three days and increasing the yield of lycopene by 500 percent,” Harris says. “Can you imagine how long it would take to generate 15 billion genetic variants with traditional cloning techniques? It would take years.”
The technique could also be used to design models of diseases — in tissue cultures, or animals — that have large-scale genomic changes.
Church said that MAGE might end up being more useful than building entire genomes from scratch. That approach is flashy and powerful, but unnecessarily complicated.