Combinatorial Evolution of DNA with RECODE
In past decades, DNA engineering protocols have led to the rapid development of synthetic biology. To engineer the natural proteins, many directed evolution methods based on molecular biology have been presented for generating genetic diversity or obtaining specific properties. Here, we provide a simple (PCR operation), efficient (larger amount of products), and powerful (multiple point mutations, deletions, insertions, and combinatorial multipoint mutagenesis) RECODE method, which is capable of reediting the target DNA flexibly to restructure regulatory regions and remodel enzymes by using the combined function of the thermostable DNA polymerase and DNA ligase in one pot. RECODE is expected to be an applicable choice to create diverse mutant libraries for rapid evolution and optimization of enzymes and synthetic pathways.
KeywordsCombinatorial multiple mutagenesis Single-stranded DNA oligonucleotides Directed evolution Pathway optimization Synthetic biology
This work was financially supported by the National Natural Science Foundation of China (31670092), the Fundamental Research Funds for the Central Universities (JUSRP51707A), and Program for Changjiang Scholars and Innovative Research Team in University (No. IRT_15R26).
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