940323 Laboratory course in molecular biology II

Type
Exercise course
Semester hours
3
Lecturer (assistant)
Organisation
Offered in
Wintersemester 2022/23
Languages of instruction
Englisch
Content

Participants will acquire basic concepts and techniques of molecular biology in two units (duration one week each).
It is also possible to complete these exercises as an individual laboratory practical course (see description "Individual course – project work").

The part "E. coli and bacteriophage lambda" should provide a deepened understanding of E. coli, the main workhorse in molecular biology, and should provide an overview about the interaction between a host cell and a virus, between E. coli and bacteriophage lambda.
In the oral entry exam (colloquium) topics like genetic nomenclature, conditional mutants and the life cycle of lambda will be discussed. Various molecular biology tools originating from the study of phages will be covered (restriction/modification, Golden Gate cloning, cosmids, Gateway cloning, protein expression with BL21/DE3, Cre-lox). Also the adaptive immunity against invasive DNA in bacteria, the CRISPR/Cas system, will be shortly discussed. In the experimental part the interaction of various phage vectors with hosts (of different genotype) will be tested and the outcome discussed in the protocol (λgt10-hflA positive selection, λgt11-supF, λEMBL3-recA). In addition, Cre-lox mediated in vivo excision of a plasmid from the phage vector λTriplEx will be performed.


"Ex. D": E. coli plasmid preparation, cultivation of mouse fibroblasts, transfection, induction of gene expression, detection of gene product (luciferase activity).

Individual course – project work:
In this case, the project work in the laboratory lasts four weeks and can also be carried out in connection with a planned master's thesis. In order to positively complete the exercises in this mode, a work plan should be prepared which - similar to the organized laboratory exercises - contains essential methods of molecular biology or molecular genetics, such as PCR, cloning, sequencing, sequence analysis, transcription analysis, transformation/transfection, Southern, SDS-PAGE, Western, heterologous protein expression, etc. For this purpose, the theoretical background should also be worked out by the students and included in the protocol. If MoBiUE-2 is completed as a project work in the laboratory, the two entry exams do not need to be passed. The group in which the project work should be carried out must be chosen and contacted by the students themselves. If there are any uncertainties or open questions, please contact the responsible course coordinator for this individual mode, Prof. Joseph Strauss (joseph.strauss@boku.ac.at).
Objective (expected results of study and acquired competences)

Upon completion of the course, the students

- are capable of using basic molecular biological tools to tackle tasks in biotechnology and genetics
- understand the advantages of phage-based gene delivery systems
- appreciate the critical factors for phage replication in bacteria
- have acquired basic knowledge of the procedures used for purification of plasmid DNA
- have gained first experience in mammalian cell culture
- know the principles governing transfection of mammalian cells
- have learned how to detect the activity of reporter gene constructs by enzyme activity asays
You can find more details like the schedule or information about exams on the course-page in BOKUonline.