title: Generation of BAC transgenic epithelial organoids.
abstract: Under previously developed culture conditions, mouse and human intestinal epithelia can be cultured and expanded over long periods. These so-called organoids recapitulate the three-dimensional architecture of the gut epithelium, and consist of all major intestinal cell types. One key advantage of these ex vivo cultures is their accessibility to live imaging. So far the establishment of transgenic fluorescent reporter organoids has required the generation of transgenic mice, a laborious and time-consuming process, which cannot be extended to human cultures. Here we present a transfection protocol that enables the generation of recombinant mouse and human reporter organoids using BAC (bacterial artificial chromosome) technology.
authors.affiliation: Hubrecht Institute, KNAW and University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Gerald
authors.initials: G
authors.lastname: Schwank
authors.forename: Amanda
authors.initials: A
authors.lastname: Andersson-Rolf
authors.forename: Bon-Kyoung
authors.initials: BK
authors.lastname: Koo
authors.forename: Nobuo
authors.initials: N
authors.lastname: Sasaki
authors.forename: Hans
authors.initials: H
authors.lastname: Clevers
mesh_terms: D000818:Animals; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics*; D004848:Epithelium / Q000254:growth & development / Q000378:metabolism*; D005818:Genetic Engineering / Q000379:methods; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006657:Histones / Q000235:genetics; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000166:cytology; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D009940:Organoids / Q000166:cytology / Q000378:metabolism*; D046509:Tissue Culture Techniques / Q000379:methods; D014162:Transfection / Q000379:methods*; D019076:Transgenes / Q000235:genetics; D014404:Tubulin / Q000235:genetics; D014741:Video Recording / Q000379:methods; D000097573:Red Fluorescent Protein
chemical_list: D049452:Green Fluorescent Proteins; D006657:Histones; D008164:Luminescent Proteins; D014404:Tubulin; D000097573:Red Fluorescent Protein