{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,15]],"date-time":"2026-09-15T12:33:10Z","timestamp":1789475590351,"version":"build-2803163510"},"reference-count":23,"publisher":"Wiley","issue":"8","license":[{"start":{"date-parts":[[2022,8,17]],"date-time":"2022-08-17T00:00:00Z","timestamp":1660694400000},"content-version":"vor","delay-in-days":16,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["U19AI144133"],"award-info":[{"award-number":["U19AI144133"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Current Protocols"],"published-print":{"date-parts":[[2022,8]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Although the isolation of <jats:italic>Treponema pallidum<\/jats:italic> subsp. <jats:italic>pallidum<\/jats:italic> (<jats:italic>T. pallidum<\/jats:italic>) from a syphilis patient dates to 1912, for the duration of the 20<jats:sup>th<\/jats:sup> century, this pathogen has remained an exceedingly difficult organism to study due to the lack of a system to support its viability <jats:italic>in vitro<\/jats:italic>. This limitation, in turn, has precluded the application of genetic engineering techniques via transformation and subsequent selection of <jats:italic>T. pallidum<\/jats:italic> transformants. A recently described method for <jats:italic>in vitro<\/jats:italic> cultivation of <jats:italic>T. pallidum<\/jats:italic>, however, has made it possible for us to experiment with transformation and selection methods. Here we describe the approach that we adopted to successfully transform <jats:italic>T. pallidum<\/jats:italic> with foreign DNA and select the resulting recombinant strain using kanamycin. \u00a9 2022 Wiley Periodicals LLC.<\/jats:p><jats:p><jats:bold>Basic Protocol 1<\/jats:bold>: Transformation of <jats:italic>T<\/jats:italic>. <jats:italic>pallidum<\/jats:italic><\/jats:p><jats:p><jats:bold>Support Protocol 1<\/jats:bold>: Quantification of <jats:italic>T. pallidum<\/jats:italic> in suspensions using dark\u2010field microscopy<\/jats:p><jats:p><jats:bold>Support Protocol 2<\/jats:bold>: Counting cells using a hemacytometer<\/jats:p><jats:p><jats:bold>Basic Protocol 2<\/jats:bold>: Selection, initial passaging, and expansion of transformed cultures<\/jats:p><jats:p><jats:bold>Basic Protocol 3<\/jats:bold>: Isolation of a clonal strain through limiting dilution<\/jats:p>","DOI":"10.1002\/cpz1.507","type":"journal-article","created":{"date-parts":[[2022,8,17]],"date-time":"2022-08-17T09:03:26Z","timestamp":1660727006000},"source":"Crossref","is-referenced-by-count":14,"title":["<i>In Vitro<\/i> Transformation and Selection of <i>Treponema pallidum<\/i> subsp. <i>pallidum<\/i>"],"prefix":"10.1002","volume":"2","author":[{"given":"Amber","family":"Phan","sequence":"first","affiliation":[{"name":"Department of Medicine, Division of Allergy and Infectious Diseases University of Washington, Harborview Medical Center  Seattle Washington"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Emily","family":"Romeis","sequence":"additional","affiliation":[{"name":"Department of Medicine, Division of Allergy and Infectious Diseases University of Washington, Harborview Medical Center  Seattle Washington"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lauren","family":"Tantalo","sequence":"additional","affiliation":[{"name":"Department of Medicine, Division of Allergy and Infectious Diseases University of Washington, Harborview Medical Center  Seattle Washington"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lorenzo","family":"Giacani","sequence":"additional","affiliation":[{"name":"Department of Medicine, Division of Allergy and Infectious Diseases University of Washington, Harborview Medical Center  Seattle Washington"},{"name":"Department of Global Health University of Washington, Harborview Medical Center  Seattle 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