Advanced Methods in Biochemistry
The course provides a thorough overview of modern biochemical and molecular biological methods, as well as available resources for gaining knowledge about these methods.
‘Methods in Biochemistry’ will introduce you to the fundamental principles behind several basic and more advanced techniques commonly used in biochemical research. During the course, the strengths and weaknesses of each technique will be covered to explain which techniques are suitable for particular applications.
Upon establishing this foundation, you will be taught how to combine different methods to overcome problems associated with the individual techniques. The theoretical aspect of the course will be supplemented with a lab practical where you will apply a number of these techniques yourself. In the end, you should be able to design strategies to address complex biological questions using the techniques you have been exposed to from a theoretical and practical perspective.
Upon completion of the course you should be able to:
- Describe the principles behind a number of common biochemical techniques.
- Explain the strengths and weaknesses of a technique for particular applications.
- Combine different biochemical methods to address a complex biological question.
- Troubleshoot biochemical methods based on their scientific principles.
- Analyze generated data and communicate them in writing and orally.
- Read, communicate and critically evaluate course-related scientific literature.
You will be expected to:
- Attend and actively participate in the lectures and tutorials.
- Participate in discussions with other students and the faculty.
- Read the assigned literature and complete the assignments and the lab report on time.
- Present the lab practical results in writing and make the appropriate corrections.
-Cell homogenization and fractionation The organization and chemical composition of pro- and eukaryotic cells will be discussed in relationship to different homogenization methods, which are used to enrich for particular fractions from these cells.
-Materials on how to make buffers and solutions (incl. sterilization), pH, the metric system, statistics and making lab. notes are provided at the beginning of the course.
-‘History of molecular biology’ Students will be taught the history behind the main organisms used for molecular biology and how these are handled, genotyped, cultured and stored.
-Centrifugation The theoretical basis for multiple centrifugation techniques will be explained as well as their applications. These include: differential centrifugation, density gradient centrifugation and analytical ultracentrifugation.
-Recombinant DNA techniques Several modern DNA analysis methods will be covered ranging from DNA isolation and sequencing to PCR and a variety of molecular biology techniques for DNA manipulation. Significant time will be spent on: restriction enzyme digests and other reactions that modify DNA, PCR, primer design, sequencing methods, homology cloning, Gibson cloning, recombineering, Cas9-based genome editing, forwarded evolution etc.
-Protein production Several different eukaryotic and prokaryotic protein production platforms are available. These will be thoroughly explained from both a theoretical and need perspective based on the current protein production bottlenecks. Specifically, we will address how to choose an organism, the expression system, and the design of the target gene.
-Protein isolation using physiochemical properties The biochemical principles for a number of basic protein purification techniques will be covered. This will include the buffer systems and resins that support: affinity chromatography, ion exchange chromatography, hydrophobic interaction chromatography, and gel-filtration. We will also go over which of these are suitable for HPLC and why.
-Antibodies and their applications We will discuss the factors to consider for generating a polyclonal/monoclonal antibody, alternative binding approaches that can replace antibodies, and several applications such as: immuno-blotting, immuno-precipitations, and ELISAs.
-Mass spectrometry Basics of mass spectrometry, MALDI-TOF, MS/MS (incl. sequencing proteins), quantitative proteomics (e.g., iTRAQ, iCAT, spectral counting), characterization of post-translational modifications using MS etc.
-Gel-electrophoresis and detection methods Different gel-electrophoresis techniques for DNA and protein will be explained as well as a number of methods for staining and detection.
-Microarrays DNA + protein arrays, RNA-seq, ribosome profiling etc.
Lectures and tutorials
All the lectures are linked to tutorials, which are a significant part of the course. For the tutorials you get assignments (‘homework’), i.e., you have to answer questions based on the lectures and articles you have to read.
The assignments are handed in, checked (feedback) and graded before the tutorial (-: fail, +: good, ++: very good). In case the tutorial is graded with a ‘fail’, the tutorial has to be corrected until it is graded with at least a ‘good’.
To be able to finish the course all tutorials should have been graded with a ‘good’ or ‘very good’.
There is one ‘special’ tutorial: the cloning tutorial. In the cloning tutorial the you design constructs (primer design, plasmid design, and you learn how to use different computer programs to facilitate this).
-Protein production practical Primer design, PCR, cloning, overexpression screening, monitoring, protein production, cell breakage, centrifugation, protein characterization (SDS-PAGE, immuno-blotting).
You write a lab. report that is corrected (until the level of the reports is acceptable).
Written exam, and the student have to complete all tutorials and lab. reports on time.
Jan-Willem de Gier
Phone: +46-8-162420 (office)
Course literatureNote that the course literature can be changed up to two months before the start of the course.
Book + course materials:
- "Principles and Techniques of Biochemistry and Molecular Biology (8thedtion)". Edited by Andreas Hofmannn and Samuel Clokie (Cambridge University Press).
- Hand-outs, articles etc. in electronic form will be provided.