636-0204-00L Lab Course: Microbial Biotechnology
Semester | Herbstsemester 2018 |
Dozierende | M. Held |
Periodizität | jährlich wiederkehrende Veranstaltung |
Lehrsprache | Englisch |
Kommentar | Only for Biotechnology MSc, Programme Regulations 2017. |
Kurzbeschreibung | Students will learn the foundations of monoseptic working practice and create and screen microbial libraries for identification of strains expressing different fluorescent protein (XFP) levels |
Lernziel | Students will learn the foundations of monoseptic working practice and create and screen microbial libraries for identification of strains expressing different fluorescent protein (XFP) levels |
Inhalt | Block A: Handling and preparation and of microbial libraries D1: Introduction to microbiological cultures and monoseptic working techniques. D2: Plasmid-based expression systems and variation of XFP synthesis levels via site-directed RBS mutagenesis. Block B: Library screening D3: In vivo screening for XFP expression levels. D4: Analysis of XFP levels via SDS-PAGE analysis. RBS-sequencing. Block C: Hit recovery and validation D5: In silico analysis of RBS variants. D6: Cellular XFP content for selected variants at different culture conditions. Block D: Data analysis and presentation D7: Protein expression analysis. Q&A for reports and presentations. D8: Final presentations and wrap-up. |
Skript | Material will be provided during the course. |
Literatur | (1) Reetz MT, Kahakeaw D, and Lohmer R. "Addressing the numbers problem in directed evolution." ChemBioChem 2008 (2) Jeschek M, Gerngross D, and Panke S. “Rationally reduced libraries for combinatorial pathway optimization minimizing experimental effort.” Nat. Commun. 2016 (3) Salis HM. “The ribosome binding site calculator.” Methods Enzymol. 2011 (4) Nienhaus G, Nienhaus K, and Wiedenmann J. "Structure–Function Relationships in Fluorescent Marker Proteins of the Green Fluorescent Protein Family." Fluorescent Proteins I. Springer Berlin Heidelberg, 2011 General introduction to microbiology: (5) Schlegel HG, and Zaborosch C. “General Microbiology.” Cambridge University Press 1993 (6) Pirt JS. “Principles of microbe and cell cultivation.” Blackwell Scientific Publications 1975 |