title: Combined chemical and physical transformation method with RbCl and sepiolite for the transformation of various bacterial species.
abstract: DNA transformation that delivers plasmid DNAs into bacterial cells is fundamental in genetic manipulation to engineer and study bacteria. Developed transformation methods to date are optimized to specific bacterial species for high efficiency. Thus, there is always a demand for simple and species-independent transformation methods. We herein describe the development of a chemico-physical transformation method that combines a rubidium chloride (RbCl)-based chemical method and sepiolite-based physical method, and report its use for the simple and efficient delivery of DNA into various bacterial species. Using this method, the best transformation efficiency for Escherichia coli DH5α was 4.3×106CFU/μg of pUC19 plasmid, which is higher than or comparable to the reported transformation efficiencies to date. This method also allowed the introduction of plasmid DNAs into Bacillus subtilis (5.7×103CFU/μg of pSEVA3b67Rb), Bacillus megaterium (2.5×103CFU/μg of pSPAsp-hp), Lactococcus lactis subsp. lactis (1.0×102CFU/μg of pTRKH3-ermGFP), and Lactococcus lactis subsp. cremoris (2.2×102CFU/μg of pMSP3535VA). Remarkably, even when the conventional chemical and physical methods failed to generate transformed cells in Bacillus sp. and Enterococcus faecalis, E. malodoratus and E. mundtii, our combined method showed a significant transformation efficiency (2.4×104, 4.5×102, 2×101, and 0.5×101CFU/μg of plasmid DNA). Based on our results, we anticipate that our simple and efficient transformation method should prove usefulness for introducing DNA into various bacterial species without complicated optimization of parameters affecting DNA entry into the cell.
authors.affiliation: School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea.
authors.forename: Jun
authors.initials: J
authors.lastname: Ren
authors.affiliation: School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea.
authors.forename: Haram
authors.initials: H
authors.lastname: Lee
authors.affiliation: Department of Chemical and Biomolecular Engineering (BK21 plus program), KAIST, Daejeon, 34141, Republic of Korea.
authors.forename: Seung Min
authors.initials: SM
authors.lastname: Yoo
authors.affiliation: School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea.
authors.forename: Myeong-Sang
authors.initials: MS
authors.lastname: Yu
authors.affiliation: School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea. Electronic address: heysoo@cau.ac.kr.
authors.forename: Hansoo
authors.initials: H
authors.lastname: Park
authors.affiliation: School of Integrative Engineering, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea. Electronic address: blisszen@cau.ac.kr.
authors.forename: Dokyun
authors.initials: D
authors.lastname: Na
mesh_terms: D001407:Bacillus / Q000235:genetics; D001410:Bacillus megaterium / Q000235:genetics; D001412:Bacillus subtilis / Q000235:genetics; D001419:Bacteria / Q000235:genetics*; D002712:Chlorides / Q000494:pharmacology*; D004269:DNA, Bacterial / Q000235:genetics*; D013293:Enterococcus faecalis / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D018014:Gene Transfer Techniques; D013294:Lactococcus lactis; D017633:Magnesium Silicates / Q000494:pharmacology*; D010957:Plasmids / Q000235:genetics; D012413:Rubidium / Q000494:pharmacology*; D014169:Transformation, Bacterial / Q000187:drug effects* / Q000235:genetics
chemical_list: D002712:Chlorides; D004269:DNA, Bacterial; D017633:Magnesium Silicates; C001671:magnesium trisilicate; D012413:Rubidium; C032710:rubidium chloride
keywords: Bacteria; Chemical transformation; Physical transformation; Rubidium chloride; Sepiolite; Transformation