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• W1966510955 abstract "Multiplex Automated Genome Engineering (MAGE) allows simultaneous mutagenesis of multiple target sites in bacterial genomes using short oligonucleotides. However, large-scale mutagenesis requires hundreds to thousands of unique oligos, which are costly to synthesize and impossible to scale-up by traditional phosphoramidite column-based approaches. Here, we describe a novel method to amplify oligos from microarray chips for direct use in MAGE to perturb thousands of genomic sites simultaneously. We demonstrated the feasibility of large-scale mutagenesis by inserting T7 promoters upstream of 2585 operons in E. coli using this method, which we call Microarray-Oligonucleotide (MO)-MAGE. The resulting mutant library was characterized by high-throughput sequencing to show that all attempted insertions were estimated to have occurred at an average frequency of 0.02% per locus with 0.4 average insertions per cell. MO-MAGE enables cost-effective large-scale targeted genome engineering that should be useful for a variety of applications in synthetic biology and metabolic engineering." @default.
• W1966510955 created "2016-06-24" @default.
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• W1966510955 date "2014-06-20" @default.
• W1966510955 modified "2023-10-12" @default.
• W1966510955 title "Direct Mutagenesis of Thousands of Genomic Targets Using Microarray-Derived Oligonucleotides" @default.
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• W1966510955 doi "https://doi.org/10.1021/sb5001565" @default.
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