Artificial Chromosomes for Disease Modeling

Credits; TheScientist

Synthetic chromosomes are ideal delivery systems for ferrying large sections of human DNA into cells. In contrast to viral vectors, human artificial chromosomes (HAC) carry more genetic material and are less likely to trigger an immune response. So far, however, technical problems have prevented HAC from reaching their full potential. Now, in a paper published in Science, researchers described an improved technique for engineering HAC that sidesteps previous barriers.

Earlier methods to synthesize HAC relied on linking shorter DNA constructs into a larger chromosome within the cell in a process called multimerization. However, the genetic fragments tended to connect in unpredictable sequences of varying lengths, making it difficult to anticipate how the genes would behave. Furthermore, the constructs often attached to natural chromosomes, potentially disrupting the host cell’s genome.

In the new study, researchers at the University of Pennsylvania circumvented this problem, known as uncontrolled multimerization, by synthesizing larger strands of DNA so that HAC could be formed from a single construct. Instead of creating 200 kilobase pair sequences, the researchers increased the size of the DNA construct to approximately 750 kilobases, removing the need for multimerization.

“We hypothesized that to [avoid] multimerization, we would have to start big,” said University of Pennsylvania biochemist Ben Black, who led the research.

Within each HAC, the researchers designed sequences to bind centromeric proteins, which ensured that daughter cells would inherit the chromosome when the original cell divided. Specifically, they added hundreds of DNA binding sites for the bacterial Lac repressor protein (LacI). By engineering the host cells to express a fusion protein—LacI fused to another protein that binds to a key centromeric protein—the team recruited proteins to form the centromere on the DNA construct.




By Holly Barker, PhD

Article can be accessed on: The Scientist