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A multifunctional system for genome editing and large-scale interspecies gene transfer.

ABSTRACT: CRISPR SWAPnDROP extends the limits of genome editing to large-scale in-vivo DNA transfer between bacterial species. Its modular platform approach facilitates species specific adaptation to confer genome editing in various species. In this study, we show the implementation of the CRISPR SWAPnDROP concept for the model organism Escherichia coli, the fast growing Vibrio natriegens and the plant pathogen Dickeya dadantii. We demonstrate the excision, transfer and integration of large chromosomal regions between E. coli, V. natriegens and D. dadantii without size-limiting intermediate DNA extraction. CRISPR SWAPnDROP also provides common genome editing approaches comprising scarless, marker-free, iterative and parallel insertions and deletions. The modular character facilitates DNA library applications, and recycling of standardized parts. Its multi-color scarless co-selection system significantly improves editing efficiency and provides visual quality controls throughout the assembly and editing process.

SUBMITTER: Teufel M

PROVIDER: S-EPMC9198041 | biostudies-literature | 2022 Jun

REPOSITORIES: biostudies-literature

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A multifunctional system for genome editing and large-scale interspecies gene transfer.

Teufel Marc M Klein Carlo A CA Mager Maurice M Sobetzko Patrick P

Nature communications 20220614 1

CRISPR SWAPnDROP extends the limits of genome editing to large-scale in-vivo DNA transfer between bacterial species. Its modular platform approach facilitates species specific adaptation to confer genome editing in various species. In this study, we show the implementation of the CRISPR SWAPnDROP concept for the model organism Escherichia coli, the fast growing Vibrio natriegens and the plant pathogen Dickeya dadantii. We demonstrate the excision, transfer and integration of large chromosomal re ...[more]

PMID: 35701417

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Project description:Lateral gene transfer (LGT) among Chlamydia trachomatis strains is common, in both isolates generated in the laboratory and those examined directly from patients. In contrast, there are very few examples of recent acquisition of DNA by any Chlamydia spp. from any other species. Interspecies LGT in this system was analyzed using crosses of tetracycline (Tc)-resistant C. trachomatis L2/434 and chloramphenicol (Cam)-resistant C. muridarum VR-123. Parental C. muridarum strains were created using a plasmid-based Himar transposition system, which led to integration of the Camr marker randomly across the chromosome. Fragments encompassing 79% of the C. muridarum chromosome were introduced into a C. trachomatis background, with the total coverage contained on 142 independent recombinant clones. Genome sequence analysis of progeny strains identified candidate recombination hot spots, a property not consistent with in vitroC. trachomatis × C. trachomatis (intraspecies) crosses. In both interspecies and intraspecies crosses, there were examples of duplications, mosaic recombination endpoints, and recombined sequences that were not linked to the selection marker. Quantitative analysis of the distribution and constitution of inserted sequences indicated that there are different constraints on interspecies LGT than on intraspecies crosses. These constraints may help explain why there is so little evidence of interspecies genetic exchange in this system, which is in contrast to very widespread intraspecies exchange in C. trachomatisIMPORTANCE Genome sequence analysis has demonstrated that there is widespread lateral gene transfer among strains within the species C. trachomatis and with other closely related Chlamydia species in laboratory experiments. This is in contrast to the complete absence of foreign DNA in the genomes of sequenced clinical C. trachomatis strains. There is no understanding of any mechanisms of genetic transfer in this important group of pathogens. In this report, we demonstrate that interspecies genetic exchange can occur but that the nature of the fragments exchanged is different than those observed in intraspecies crosses. We also generated a large hybrid strain library that can be exploited to examine important aspects of chlamydial disease.

Dataset Information

A multifunctional system for genome editing and large-scale interspecies gene transfer.

Publications

A multifunctional system for genome editing and large-scale interspecies gene transfer.

Similar Datasets

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