Publikationsdatenbank

Genetic modification of bovine intestinal epithelial cells for interaction studies with zoonotic pathogens in 3D-organoids (2019)

Art

Poster

Autoren

Sharbati, Soroush (WE 3)

Häger, Jan-Dirk

Menge, Christian

Berens, Christian

Kongress

Zoonoses 2019 - International Symposium on Zoonoses Research

Berlin, 16. – 18.10.2019

Quelle

Zoonoses 2019 - International Symposium on Zoonoses Research : Book of Abstracts — International Symposium on Zoonoses Research (Hrsg.)

Berlin, 2019 — S. 213

Sprache

Englisch

Verweise

URL (Volltext): https://evis.events/event/79/attachments/23/154/Book_of_Abstracts_Zoonoses2019.pdf

Kontakt

Institut für Veterinär-Biochemie

Oertzenweg 19 b
14163 Berlin
+49 30 838 62225
biochemie@vetmed.fu-berlin.de

Abstract / Zusammenfassung

Withinthe framework of the pilot project '3D-KOR' funded by the National Research Platform for Zoonoses, new bovine host-microbiota interaction models are established. The aim of the project is the development of an intestinal 3D cell culture system from bovine colon. To foster the generation of 3D-organoids in vitro,primary intestinal crypt epithelial cells (pCEC) and de-differentiated cell lines (FKD-R) are genetically edited.Immortalisation or re-differentiation is realised by CRISPR/Cas9-mediatedgenome editing using TERT (telomerase) and miR-147b (Sharbati et al., 2015), respectively. Both the immortalisation of pCEC and the re-differentiation of FKD-R are achieved by integrating an expression cassette (EC) containing TERT or miR-147b in a 'safe harbour locus' (Wu et al., 2015). It is integrated into the bovine genome by CRISPR/Cas9 and homology directed repair. For the universal generation of the required repair template, we have developed a highly efficient assembly PCR which providesan EC within a few days. ECs produced this way are currently used and suitable clones are characterised. These models provide a versatile basis for later complex studies dealing with interactions of zoonotic pathogens (e.g. Shiga toxin-producing E.coli,Campylobacter, Shigella or Mycobacteria) or components of the microbiota with the bovine intestine.