SURE editing: combining oligo-recombineering and programmable insertion/deletion of selection markers to efficiently edit the Mycoplasma pneumoniae genome
Author
Eva, Garcia-Ramallo
Miravet-Verde, Samuel
Burgos, Raul
Scarpa, Margherita
Serrano, Luis
Lluch-Senar, Maria
Publication date
2022ISSN
0305-1048
Abstract
The development of advanced genetic tools is boosting microbial engineering which can potentially tackle wide-ranging challenges currently faced by our society. Here we present SURE editing, a multi-recombinase engineering rationale combining oligonucleotide recombineering with the selective capacity of antibiotic resistance via transient insertion of selector plasmids. We test this method in Mycoplasma pneumoniae, a bacterium with a very inefficient native recombination machinery. Using SURE editing, we can seamlessly generate, in a single step, a wide variety of genome modifications at high efficiencies, including the largest possible deletion of this genome (30 Kb) and the targeted complementation of essential genes in the deletion of a region of interest. Additional steps can be taken to remove the selector plasmid from the edited area, to obtain markerless or even scarless edits. Of note, SURE editing is compatible with different site-specific recombinases for mediating transient plasmid integration. This battery of selector plasmids can be used to select different edits, regardless of the target sequence, which significantly reduces the cloning load associated to genome engineering projects. Given the proven functionality in several microorganisms of the machinery behind the SURE editing logic, this method is likely to represent a valuable advance for the synthetic biology field.
Document Type
Article
Document version
Published version
Language
English
Subject (CDU)
57 - Biological sciences in general
Keywords
Biologia sintètica
Clonació de muntatges
Recombinació
Biología sintética
Clonación de ensamblaje
Recombinación
Synthetic Biology
Assembly Cloning
Recombination
Pages
19
Publisher
Oxford University Press
Is part of
Nucleic Acids Research
Citation
Piñero Lambea, Carlos; Eva, Garcia-Ramallo; Miravet-Verde, Samuel [et al.]. SURE editing: combining oligo-recombineering and programmable insertion/deletion of selection markers to efficiently edit the Mycoplasma pneumoniae genome. Nucleic Acids Research, 2022, [p. 1-19]. Disponible en: <https://academic.oup.com/nar/advance-article/doi/10.1093/nar/gkac836/6754916?login=true>. Fecha de acceso: 2 nov. 2022. DOI: 10.1093/nar/gkac836
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- Ciències Bàsiques [65]
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