Improving the Transformation Efficiency of Synechococcus sp. PCC 7002 via Methylome-Guided Premethylation of DNA

Hren, Andrew; Alexander, William G.; Abraham, Joshua P.; Tumen-Velasquez, Melissa P.; Vergara, Michael Melesse; Guss, Adam M.; Pfleger, Brian F.; Fox, Jerome M.; Eckert, Carrie A.

doi:10.1021/acssynbio.5c00370

Title: Improving the Transformation Efficiency of Synechococcus sp. PCC 7002 via Methylome-Guided Premethylation of DNA

Journal Article · Mon Aug 04 00:00:00 UTC 2025 · ACS Synthetic Biology

DOI: https://doi.org/10.1021/acssynbio.5c00370 · OSTI ID:2575345

Hren, Andrew ^[1];

^[2];

^[3]; Tumen-Velasquez, Melissa P. ^[2];

^[2];

^[2]; Pfleger, Brian F. ^[3];

^[1];

^[2]

Univ. of Colorado, Boulder, CO (United States)
Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)
Univ. of Wisconsin, Madison, WI (United States)

Cyanobacteria are promising microbial platforms for a diverse set of biotechnology applications, from living materials to photosynthetic chemical production, but are less well characterized than commonly engineered microbes such as Escherichia coli. This study facilitates genetic engineering in Synechococcus sp. PCC 7002, a fast-growing, halotolerant, and naturally competent strain, by identifying ten native methylation motifs and designing shuttle strains that mimic the native methylation state by expressing a subset of heterologous methyltransferases. DNA methylation in E. coli with as few as two active methyltransferases increased transformation efficiency up to 30-fold across four distinct integration sites in PCC 7002. This work provides an experimental framework to bypass native restriction-modification systems for efficient genome editing and metabolic engineering in nonmodel bacteria.

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Research Organization:: Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Biological and Environmental Research (BER)

Grant/Contract Number:: AC05-00OR22725; SC0018368; SC0019404

OSTI ID:: 2575345

Journal Information:: ACS Synthetic Biology, Journal Name: ACS Synthetic Biology Journal Issue: 8 Vol. 14; ISSN 2161-5063

Publisher:: American Chemical Society (ACS)Copyright Statement

Country of Publication:: United States

Language:: English

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Related Subjects

59 BASIC BIOLOGICAL SCIENCES
Bacteria
DNA methylation
Genetics
Genomics
Organic reactions
Peptides and proteins
cyanobacteria
genetic engineering
restriction modification
transformation efficiency

Title: Improving the Transformation Efficiency of Synechococcus sp. PCC 7002 via Methylome-Guided Premethylation of DNA

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References (32)

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