VEGAS as a Platform for Facile Directed Evolution in Mammalian Cells
| Autor: | Justin G. English, Bryan L. Roth, Reid H.J. Olsen, Ryan T. Strachan, Adam S. Cockrell, Daniel Wacker, Michael T Patel, Darshan Singh, Karoline White, Katherine Lansu |
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| Rok vydání: | 2018 |
| Předmět: |
Genetic Vectors
Computational biology Alphavirus Biology General Biochemistry Genetics and Molecular Biology Article Receptors G-Protein-Coupled 03 medical and health sciences 0302 clinical medicine Allosteric Regulation Fluorescence Resonance Energy Transfer Animals Humans Vector (molecular biology) Amino Acid Sequence Transcription factor Selection (genetic algorithm) 030304 developmental biology 0303 health sciences RNA Single-Domain Antibodies biology.organism_classification Directed evolution HEK293 Cells Viral evolution Mutation Sindbis Virus Directed Molecular Evolution Sequence Alignment 030217 neurology & neurosurgery Transcription Factors |
| Zdroj: | Cell |
| ISSN: | 1097-4172 |
| Popis: | Summary Directed evolution, artificial selection toward designed objectives, is routinely used to develop new molecular tools and therapeutics. Successful directed molecular evolution campaigns repeatedly test diverse sequences with a designed selective pressure. Unicellular organisms and their viral pathogens are exceptional for this purpose and have been used for decades. However, many desirable targets of directed evolution perform poorly or unnaturally in unicellular backgrounds. Here, we present a system for facile directed evolution in mammalian cells. Using the RNA alphavirus Sindbis as a vector for heredity and diversity, we achieved 24-h selection cycles surpassing 10−3 mutations per base. Selection is achieved through genetically actuated sequences internal to the host cell, thus the system’s name: viral evolution of genetically actuating sequences, or “VEGAS.” Using VEGAS, we evolve transcription factors, GPCRs, and allosteric nanobodies toward functional signaling endpoints each in less than 1 weeks’ time. |
| Databáze: | OpenAIRE |
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