The design and in vitro synthesis of genes or long DNA molecules is an important and powerful tool in biomedical research, synthetic biology and protein engineering. While the actual synthesis and assembly reactions are highly amenable to automation and parallel processing, the major bottlenecks for making in vitro DNA synthesis a routine, high throughput and high speed process are the verification and recovery of the full length and error free synthetic DNA molecules which rely on laborious E. coli cloning and Sanger sequencing technologies. We propose the development of a bench top microfluidic system that has the ability to perform DNA synthesis, sequence verification and recovery of the desired full length DNA molecules. The system incorporates the innovative and flexible digital microfluidic platform developed by Advanced Liquid Logic, Inc. (ALL) with an efficient gene assembly protocol developed by the J. Craig Venter Institute (JCVI). The resulting system will employ pyrosequencing chemistry on a digital microfluidic cartridge to enable sequence verification and recovery of full length, error- free DNA molecules bypassing the laborious cloning and capillary based sequencing processes. The system will also enable direct coupling of the microchip-based oligonucleotide synthesis platform to the downstream assembly, sequencing and recovery process, further reducing the costs and improving the overall efficiency of gene and genome synthesis.
(provided by applicant): The design and in vitro synthesis of genes or long DNA molecules is an important and powerful tool in biomedical research, synthetic biology and protein engineering. We propose the development of a bench top microfluidic system that has the ability to perform DNA synthesis, sequence verification and recovery of the desired full length DNA molecules.
| Khilko, Yuliya; Weyman, Philip D; Glass, John I et al. (2018) DNA assembly with error correction on a droplet digital microfluidics platform. BMC Biotechnol 18:37 |
