Molecular Biology and Gene Technology I

Data

Official data in SubjectManager for the following academic year: 2023-2024

Topic

Course lectures present the theoretic background and practical utilization of current and future-proof methodology of molecular biology, and within that genetic engineering. The course includes the following methodologies: various PCR types, restriction endonuclease digestion, molecular cloning, RNA interference and CRISPR/Cas9, transduction and transfection of pro- and eukaryotic cells, expression systems, development of genetically modified cells and organisms, gene therapy applications. During course seminars the students will familiarize with basic molecular biology and within that genetic engineering-related trouble-shooting. During course practices pipetting and precision skills, sterile techniques, master-mix preparation, serial dilutions etc. will be practiced. Solving simple practical issues will help to gain deep understanding and practical knowledge. Having fulfilled this molecular biology and genetic engineering-based course the graduates will get hands-on- expertise that will aid their efforts on the labor-market for both academic and industrial positions.

Lectures

• 1. DNA, gene structure - Dr. Kvell Krisztián
• 2. Transcription, translation, central dogma - Dr. Kvell Krisztián
• 3. Genome organisation (prokaryotes, eukaryotes) - Dr. Kvell Krisztián
• 4. RNA species focusing on mRNA and miRNA - Dr. Kvell Krisztián
• 5. Extracellular vesicle subtypes - Dr. Kvell Krisztián
• 6. Extracellular vesicles as carriers - Dr. Kvell Krisztián
• 7. Lipid nanoparticles as carriers - Dr. Kvell Krisztián
• 8. Enzyme tools: restriction endonucleases - Dr. Kvell Krisztián
• 9. Enzyme tools: ligase, topoisomerase, klenow - Dr. Kvell Krisztián
• 10. Enzyme tools: reverse transcriptase - Dr. Kvell Krisztián
• 11. Enzyme tools: heatstable polymerases - Dr. Kvell Krisztián
• 12. PCR: conventional - Dr. Kvell Krisztián
• 13. PCR: quantitative - Dr. Kvell Krisztián
• 14. PCR: absolute - Dr. Kvell Krisztián
• 15. Consultation - Dr. Kvell Krisztián
• 16. Midterm evaluation - Dr. Kvell Krisztián
• 17. Amplification-based high-throughput assay (Quantstudio) - Dr. Kvell Krisztián
• 18. Amplification-free high-throughput assay (Nanostring) - Dr. Kvell Krisztián
• 19. Plasmids in nature - Dr. Kvell Krisztián
• 20. Plasmids in the lab - Dr. Kvell Krisztián
• 21. Phages in nature - Dr. Kvell Krisztián
• 22. Phages in the lab - Dr. Kvell Krisztián
• 23. Bacterial transformation: calcium-phosphate - Dr. Kvell Krisztián
• 24. Bacterial transformation: electroporation - Dr. Kvell Krisztián
• 25. Bacterial antibiotic resistance in nature - Dr. Kvell Krisztián
• 26. Bacterial antibiotic resistance in the lab - Dr. Kvell Krisztián
• 27. Consultation - Dr. Kvell Krisztián
• 28. Final evaluation - Dr. Kvell Krisztián

Practices

• 1. DNA isolation with silica membrane I
• 2. DNA isolation with silica membrane II
• 3. RNA isolation with silica membrane I
• 4. RNA isolation with silica membrane II
• 5. Extracellular vesicle enrichment
• 6. Extracellular vesicle analysis with Nanosight
• 7. Lipid nanoparticle production with Lipofectin I
• 8. Lipid nanoparticle production with Lipofectin II
• 9. Reverse transcription using RT enzyme I
• 10. Reverse transcription using RT enzyme II
• 11. Preparing conventional PCR
• 12. Evaluating conventional PCR
• 13. Preparing quantitative PCR
• 14. Evaluating quantitative PCR
• 15. Consultation
• 16. Consultation
• 17. Evaluating Quantstudio results
• 18. Evaluating Nanostring results
• 19. Starting bacterial cultures with antibiotics
• 20. Evaluating bacterial cultures with antibiotics
• 21. Starting bacterial cultures with phages
• 22. Harvesting bacterial cultures with phages
• 23. Heat-shock transformation of bacterial cultures
• 24. Plating transformed bacteria, colony evaluation
• 25. Plasmid isolation with silica membrane I
• 26. Plasmid isolation with silica membrane II
• 27. Consultation
• 28. Consultation

Seminars

• 1. Concept of kit-based DNA isolation
• 2. Concept of kit-based RNA isolation
• 3. Concept of extracellular vesicle analysis
• 4. Concept of lipid nanoparticle production
• 5. Planning the use of enzymes in the lab
• 6. Planning a conventional PCR
• 7. Planning a quantitative PCR
• 8. Consultation
• 9. Planning the use of high-throughput assays
• 10. Planning an antibiogram
• 11. Planning bacterial phage transformation
• 12. Planning bacterial plasmid transformation
• 13. Concept of plasmid isolation
• 14. Consultation

Reading material

Obligatory literature

Cox, Doudna, O’Donnel: Molecular Biology: Principles and Practice, Freeman , 2nd Edition, 2015, ISBN:1464126143
Mitra, Sandhya: Genetic Engineering: Principles and Practice, MC GRAW HILL INDIA, 2015, ISBN-10 : 9789339203535, ISBN-13 : 978-9339203535

Literature developed by the Department

Notes

Recommended literature

Krebs, Goldstein, Kilpatrick: Lewin’s Essential Genes, Jones and Bartlett, 4th Edition, 2020, ISBN:1284173135

Conditions for acceptance of the semester

Maximum of 15 % absence allowed

Mid-term exams

Single choice questions prepared according to the subject of the lectures. The written exam test might contain minimum requirement questions where the wrong answers could mean failure on the exam.

Making up for missed classes

Not possible.

Exam topics/questions

https://gytk.pte.hu/en/menupont/10537

Examiners

• Dr. Kvell Krisztián

Instructor / tutor of practices and seminars

• Dr. Kvell Krisztián
• Garai Kitti