| Course title | Transient and stable Plant Transformation |
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| Course code | KBI/TRA |
| Organizational form of instruction | Exercise + Seminar |
| Level of course | Bachelor |
| Year of study | 3 |
| Semester | Summer |
| Number of ECTS credits | 4 |
| Language of instruction | Czech |
| Status of course | Compulsory |
| Form of instruction | Face-to-face |
| Work placements | This is not an internship |
| Recommended optional programme components | None |
| Lecturer(s) |
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| Course content |
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Content of the seminar: 1) Definition of basic concepts, history of transformation and transgenosis of plants, bacteria Agrobacterium tumefaciens - Ti plasmid. Distribution of scientific papers to students for journal clubs. 2) The principle of transformation using A. tumefaciens: the transfer of T -DNA into the plant. Other species of the Agrobacterium genus. 3) Stable vs transient transformation of plants: transgenosis mono- and dicotyledonous plants, methods of transient transformation of leaves, cotyledons, protoplasts. Methods for stable transformation of plants. 4) Components of transgenes - promotors, terminators, polyadenylation signal, 5'not coding region. 5) The gene silencing - a T-DNA insertional mutagenesis, RNAi technique. 6 ) Reporter genes and marker lines. Selection markers for plant transgenes - antibiotics, herbicides, carbohydrate source and plant hormones. 7 ) Loss of activity of transgenes - the transcriptional and post-transcriptional inactivation of the transgene following Journal club I 9) GMO - tolerance to herbicides and insect pests, protease inhibitors. Journal Club II 10) GMO - other transgenes used nowadays. GMO field research. Journal club III 11) GMO legislation in our country. GMO vs Eko -Bio sustainable agriculture, final discussion Journal Club IV 12) Final test Content of the practical seminars: 1) Arabidopsis thaliana handling. Practical part: Arabidopsis thaliana sawing - Columbia ecotype for stable floral-dip transformation. 2) E. coli a A. tumefaciens cultures, content of the media Practical part: preparation of medias for A. tumefaciens, inoculation of A. tumefaciens to the liquid and solid media 3) Transient transformation - methods overview Practical part: transient transformation of N. bentamiana leaves with A. tumefaciens 4) Practical part: microscopical observation of transformed N. benthamiana leaves 5) Practical part: transient transformation of A. thaliana seedlings with A. tumefaciens 6) Practical part: microscopical observation of transformed A. thaliana seedlings 7) Methods for bacterial DNA isolation Practical part: isolation of bacterial DNA. 8) Stable "Floral-Dip" transformation of A. thaliana plants. Practical part: stable "Floral-Dip" transformation of A. thaliana plants (prepared during the first practical seminar). 9) Practical part: A. thaliana seeds handling: AraSYSTEM. 10) Selection markers for plant transgenes. Practical part: preparation of selection media for A. thaliana seeds - sterile handling and sowing of transformed seeds. 11) Practical part: selection of transformed plants 12) Final discussion and protocols corrections.
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| Learning activities and teaching methods |
Monologic Lecture(Interpretation, Training), Dialogic Lecture (Discussion, Dialog, Brainstorming), Observation, Demonstration, Training in job and motor Skils, Laboratory Work
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| Learning outcomes |
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The aim of the seminar is to introduce following topics and themes: Basic of the plant transgenosis - transient and stable transformation , the principle of the transformation of plant cells using Agrobacterium, direct transgenosis using DNA. Application of plant transgenosis in basic research - targeted alteration of plants in relation to the study of previously uncharacterized genes, increase or downregulation of the expression of the genes of interest, T-DNA insertion mutagenesis, RNAi techniques, reporter genes, selectable markers. Application of plant transgenosis in applied research - targeted transgenosis in breeding, GMO plants (perspective transgenes integrated pest protease inhibitors), the legal regulation of GMOs in Laws and legislation . During the seminar, students will elaborate and actively present scientific articles in the form of journal clubs. During the practical seminar, students will familiarise the method of stable transformation of Arabidopsis thaliana using the "floral dip" method and with methods for transient transformation of Nicotiana bentamiana. Students will learn how to work practically with the model plant Arabidopsis thaliana.
After completing the course the student should be able to manage the most used transient and stable transformation methods used in plant biotechnology. |
| Prerequisites |
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unspecified
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| Assessment methods and criteria |
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Seminar Work, Written exam
Participation in practical exercises (100%), presentations of assigned publications and active participation in Journal club discussions, preparation of protocols from practical courses and successful passing of written test. |
| Recommended literature |
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| Study plans that include the course |
| Faculty | Study plan (Version) | Category of Branch/Specialization | Recommended semester | |
|---|---|---|---|---|
| Faculty: Faculty of Science | Study plan (Version): Biotechnology and Genetic Engineering (2022) | Category: Chemistry courses | 3 | Recommended year of study:3, Recommended semester: Summer |