Assembly of large synthetic genomes using nanofluidics

Objective

Synthetic genomics is a rapidly growing branch of synthetic biology where the goal is to assemble a complete genome from small synthetic DNA fragments. The main obstacle is the later stages of assembly where fragments on the kilobasepair length scale are to be assembled, where the efficiency rapidly falls off with increasing fragment size. A large contribution to this bottleneck is purely physical, where the probability of DNA ends meeting decreases with increasing DNA size. My solution to this problem is to force DNA ends to meet using nanofluidic channels. In such channels DNA is stretched out to almost the complete contour length, exposing the ends for hybridization. As part of my ongoing ERC-CoG, I have demonstrated several principles that will make it possible to use nanofluidic devices for genome assembly. Long DNA can be entropically trapped in nanochannels to force DNA ends to meet to promote hybridization of complementary overhangs. The bacterial protein Ku stably anneals 4 bp complementary overhangs, making it possible to engineer which fragments that are annealed. Bacterial Ligase D then binds to Ku to ligate the DNA. Within this PoC project I will take the next steps to bring this principle to a commercially feasible platform.