sb9b00322_si_001.pdf (1.75 MB)
A Toolkit for Rapid Modular Construction of Biological Circuits in Mammalian Cells
journal contribution
posted on 2019-11-04, 23:44 authored by João
Pedro Fonseca, Alain R. Bonny, G. Renuka Kumar, Andrew H. Ng, Jason Town, Qiu Chang Wu, Elham Aslankoohi, Susan Y. Chen, Galen Dods, Patrick Harrigan, Lindsey C. Osimiri, Amy L. Kistler, Hana El-SamadThe ability to rapidly assemble and
prototype cellular circuits
is vital for biological research and its applications in biotechnology
and medicine. Current methods for the assembly of mammalian DNA circuits
are laborious, slow, and expensive. Here we present the Mammalian
ToolKit (MTK), a Golden Gate-based cloning toolkit for fast, reproducible,
and versatile assembly of large DNA vectors and their implementation
in mammalian models. The MTK consists of a curated library of characterized,
modular parts that can be assembled into transcriptional units and
further weaved into complex circuits. We showcase the capabilities
of the MTK by using it to generate single-integration landing pads,
create and deliver libraries of protein variants and sgRNAs, and iterate
through dCas9-based prototype circuits. As a biological proof of concept,
we demonstrate how the MTK can speed the generation of noninfectious
viral circuits to enable rapid testing of pharmacological inhibitors
of emerging viruses that pose a major threat to human health.