Advanced techniques in molecular biotechnology 1400-226ZTBTM-en
The course focuses on current issues in molecular biotechnology. It is composed of 3 thematic blocks:
1. Genome editing by CRISPR
Basics of genome editing, CRISPR elements, history of CRISPR editing invention, application of CRISPR as molecular biology tool. Molecular aspects of CRISPR. Aplicative variants of different research models (plants, animals, microorganisms, and tissue culture) Students will design their own CRISPR/Cas experiment and perform it during the first 5 classes to finally verify the obtained clones. Students will perform the following practical tasks: primer design, guide RNA cloning, analysis of obtained clones and sequencing results, yeast cell transformation, and phenotyping by diagnostic PCR.
2. Protein as products
-Students will learn the basics about plant stress (biotic and abiotic) and will know the methods of plant genome modification to modulate a stress response. The practical part will focus on assessing the susceptibility to stress in Arabidopsis thaliana and plants used in agriculture. Stress will be induced by flagellin mimicking interaction with the pathogen and assessed by the level of reactive oxygen species in the leaf. As a final activity, students will propose a project to obtain plant variety with improved resistance to stress using genome editing techniques. The project will be prepared in small teams of 2-4 people, based on the knowledge gained during the classes and literature data. The project principles will be presented during the classes as a 10-minute short talk and delivered to the teacher in written form (5 pages maximum).
-High-throughput gene cloning using the no-ligation SLIC method as an example. Design of custom constructs and primers for cloning.
-Metagenomes as a source of new enzyme activities. Sequencing of metagenomes and laccases (oxidases) as examples of enzymes with biotechnological potential.
-Basics of protein purification and activity assessment. Students will purify recombinant proteins produced in bacteria (various new laccases found in bacterial metagenomes and other nucleic acid-binding proteins) and then assess the quality and efficiency of the purification by an SDS-PAGE analysis. They then check the activity of the proteins using a colorimetric method and determine the optimum pH of the reaction. The activity of nucleic acid-binding proteins will be asses by immunostaining of fixed cells followed by state-of-the-art microscopy and analysis of the images collected.
3. Intellectual property in biotechnology.
Students will know different forms of intellectual property, especially this protected in biotechnology,(as well as agriculture and medicine). Students will have the skills to use patent databases as a source of intellectual property knowledge and also as an alternative to scientific papers. Students will analyse the patent application, how it is proceeded, and what is the patent protection. Students will gain knowledge about biological material repositories. As a summary, the student will prepare a short presentation on the patent of choice and present it to the other participants in the course.
The exercises will take place in the classroom, interchangeably, the lectures and the experimental parts. During the course, special attention will be given to the student's individual work or to working in small teams. Communication and individual performance according to a given protocol is essential during classes (simple mathematical calculations, manual skills).
The course is designed for students who plan to work in the research and development of biotechnological companies and for those who plan a scientific career.
Main fields of studies for MISMaP
biotechnology
Type of course
Course coordinators
Learning outcomes
In terms of knowledge, students should:
-Possess advanced knowledge in the field of molecular biology and biotechnology (K_W01 Bt2).
-Understand the impact of molecular biology on the development of biotechnological solutions and know how to use it in practice(K_W03 Bt2).
Describe the mechanisms of defence of prokaryotic organisms, which became the prototype of CRISPR editing. Knows the CRISPR elements function during an editing event (K_W03 Bt2).
-Be able to use basic vocabulary related to biotechnology and molecular biology in English (K_W12 Bt2).
-Understand the importance of experimental work in biotechnology and the role of molecular analysis in biological and medical research (K_W05 Bt2).
In terms of skills, students should:
- be able to use advanced research techniques appropriate for biotechnology (K_U01Bt2)
- Be able to use basic methods of nucleic acid in silico analysis, apply the theoretical knowledge in this area (searching databases and professional literature) and analyse selected data (K_U17 Bt2).
-be able to plan experiments of heterologous protein expression, understand the applicability of such activities, and assess the chances of success of such attempts. Students are able to choose the cloning technique proposed by the lecturer (SLIC, GATEWAY, GOLDEN GATE, shotgun cloning, classic cloning using restriction enzymes), isolate nucleic acids, or analyse the sequencing results (K_U04 Bt2)..
- be able to plan experiments using CRISPR, design RNA guide and repair templates, select the method of introducing the DNA, repair template, and Cas9 nuclease into the cell. Students should be able to carry out a CRISPR experiment and verify the results of this experiment under supervision (K_U13 Bt2).
- be able to isolate and purify proteins as well as obtain biologically active preparations that have potential biotechnological applications. Perform tests of the activity of purified proteins to verify their usefulness (e.g. recombinant antibodies, restriction enzymes, protein-modifying enzymes) (K_U14 Bt2).
- be able to assess the risk of using CRISPR/Cas9 editing for genome modification and successfully counteract this phenomenon (K_U015 Bt2).
-be able to perform correct results interpretation and conclusion. Apply mathematical and statistical methods to describe phenomena and data analysis (K_U05 Bt2).
-be able to present scientific results and professional communication verbally (K_U08 Bt2).
-be able to work in a team and manage the teamwork of a small group (K_U10 Bt2)
Has the ability to use English to read scientific literature and communicate with foreigners (K_U02 Bt2).
In terms of social competences, students should:
-Understand the physical and chemical phenomena that occur in the environment (K_K01Bt2).
- Be responsible for their own work and equipment; demonstrate respect for others work (K_K03 Bt2).
- use objective sources of scientific information and draw critical conclusions while analysing and solving practical problems (K_K04 Bt2).
Assessment criteria
The final grade depends on the result of the exam (passing is the prerequisite) and other activities performed during the class.
Pass criteria:
- attendance (two absences per semester are allowed)
- preparation of a short 10-minute presentation about intellectual property (in a team of two students) will require literature data and the knowledge gained during the classes. Performance during the presentation will have an impact on the final grade;
- preparation of a short 10-minute presentation and a final report (5 pages of A4 format maximum), about protein as a product part, performed in a group of 2-4 persons, will require literature data and the knowledge gained during the course. Performance during the presentation and the report will have an impact on the final grade;
- homeworks prepared individually or in teams (mainly the design of the primers used during the course for cloning and genome editing),
- a written exam consisting mainly of closed questions and/or 3 open questions (a final score of 51% is required to pass)
Bibliography
The ATMB Handbook (2019) theoretical introduction and practical protocols prepared by the teachers of the class. A PDF file or printed version will be provided for each participant of the classes.
Genome Editing ed. Kursad Turksen, 2016
Notes
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Term 2023Z:
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Term 2024Z:
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Additional information
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