Practical Course on Rapid Cloning, Protein Expression, and Purification CPEP 18 - 3 ECTS
This course is organised in the collaboration with the Protein Expertise Platform (PEP). It is based on the EMBO PEPC5 course.
Date: Wednesday 28th February to Friday 9th March 2018
Start: Wednesday, 28th February 2018, 12:30
END: Friday, 9 March 2018, 12:00
Place: Biochemistry teaching lab, KB.F4.04, KBC-building 4th floor (2 stairs up), Linnaeus väg 6, Umeå University
Time: About 6-8 h per day, mainly laboratory work but also lectures, discussions & seminars
Course leader: Uwe Sauer, KBC, 786-5930, E-mail: firstname.lastname@example.org Instructor: Gunter Stier, Heidelberg Univ., Germany, E-mail: email@example.com
EXTENDED Registration deadline: Friday, 16th of February, 2018
(Admission on a first-come first-served basis)
The registration is closed now!
Participants: max. 12 (PhD students, post-docs or technical staff)
NO Participation fee!
Credits: 3 ECTS on a pass – no pass basis. Full time participation is required to obtain a pass certificate.
More information about the course:http://babel.ucmp.umu.se/cpep
Aims of the practical CPEP course:
After the course you should know how to:
- Plan and successfully execute your project. Bring your own project(s)!(*)
- Make efficient use of bioinformatics tools for planning and analysis of protein expression projects
- Design PCR primers and codon optimize genes
- Apply various cloning and mutagenesis techniques
- Use medium throughput approaches for cloning and protein expression in E. coli
- Carry out fast parallel cloning of your gene(s) into vectors containing different tags and fusion proteins
- Use of fusion proteins and associated purification strategies
- Test for optimal protein production: different strains, growth conditions and fusion partners
- Co-express components of protein complexes and their purification strategies
- Prepare and use of auto-inducing medium
- Do protein expression mini-screens
- Scale-up positive screening candidates to a protein midi-preparation using purification machines
- Fc-fusion protein expression in mammalian cells
- Trouble shoot problems and apply strategies to solve them
- Apply special protein labeling techniques, such as SeMet, 15N-labeling for quick NMR analysis
Additional demonstrations will include the use of:
- Protein purification with “Äkta” machines
- The Mosquito crystallization robot, crystal imaging facility
- Crystallization and X-ray diffraction of crystals and analysis of diffraction data
- NMR characterization of 15N labeled proteins and interpretation of simple NMR spectra
- EM facility – first approach to sample preparation (negative staining) and electron microscopy measurements
Practical sessions are complemented with lectures, round-table discussions about techniques, problems and their solutions. Participants will present short seminar talks of selected topics.
(*) If you bring your own project(s), you should contact the instructors at least 2 weeks before the start of the course (preferably no later than February 16, 2018).
The course leaders will provide you with a project if you do not bring one.