RECOMBINANT DNA TECHNOLOGIES FOR GENE THERAPY

1. Knowledge and Understanding

Students will acquire advanced knowledge of the principles and methodologies of recombinant DNA technology and its applications in gene therapy. In particular, they will understand gene cloning strategies, the design of recombinant DNA constructs, the main viral and non-viral gene delivery systems, and modern genome editing technologies.

2. Applying Knowledge and Understanding

Students will be able to apply the acquired knowledge to understand and theoretically design gene delivery and genome modification strategies, identifying the most appropriate molecular approaches and systems according to specific experimental and therapeutic objectives.

3. Making Judgements

Students will develop the ability to critically evaluate the potential, advantages, and limitations of different gene delivery and genome editing technologies, through the analysis of scientific literature and the comparison of different therapeutic strategies.

4. Communication Skills

Students will acquire the ability to describe and discuss the principles and applications of recombinant DNA technology and gene therapy using appropriate scientific terminology. Through the preparation and presentation of seminars, they will also develop skills in scientific communication and critical discussion of research findings.

5. Learning Skills

Students will develop the skills necessary to independently expand their knowledge of recombinant DNA technology and gene therapy through the consultation and critical interpretation of scientific literature, keeping pace with ongoing technological advances and emerging applications in the field.

The course consists of lectures complemented by the presentation and discussion of scientific articles and student-led seminars. Teaching activities are designed to promote understanding of the course topics, critical thinking, and active student participation.

Attendance is mandatory in accordance with the regulations of the Degree Programme.

Basic knowledge of molecular biology, genetics, and biochemistry is required, particularly regarding the structure and function of nucleic acids, the mechanisms of DNA replication, transcription and translation, and the regulation of gene expression. No specific prior knowledge of recombinant DNA technology or gene therapy is required.

Attendance is mandatory in accordance with the regulations of the Degree Programme.

1. Recombinant DNA Technology and Gene Cloning
Principles of recombinant DNA technology. Cloning and expression vectors. Gene cloning strategies and construction of recombinant DNA molecules. Transformation, selection, and identification of recombinant clones. Design of recombinant constructs and expression cassettes for eukaryotic cells.

2. Principles and Strategies of Gene Therapy
Molecular foundations of gene therapy. In vivo and ex vivo gene therapy. Strategies for gene introduction, replacement, and modulation of gene expression. Coding and non-coding therapeutic nucleic acids. Persistence and regulation of the expression of introduced genetic material.

3. Non-Viral Gene Delivery Systems
Major biological barriers to nucleic acid delivery. Physical and chemical methods of gene transfer. Lipid-, polymer-, and nanoparticle-based delivery systems. Advantages and limitations of non-viral approaches.

4. Viral Vectors for Gene Therapy
Principles of viral vector engineering. Retroviral and lentiviral, adenoviral, and adeno-associated viral (AAV) vectors. Molecular characteristics, production strategies, and gene transfer mechanisms. Tropism, genomic integration, and persistence of transgene expression. Advantages and limitations of major viral vector systems.

5. Genome Editing Technologies
Principles of targeted genome modification. Cellular DNA repair mechanisms. Programmable nucleases and CRISPR/Cas systems. Gene disruption and correction strategies. Base editing and prime editing technologies. Applications and limitations of genome editing approaches.

6. Applications and Future Perspectives of Gene Therapy
Major applications of gene therapy in inherited and acquired diseases. Stem cell gene therapy and cellular immunotherapy. Safety concerns, immunogenicity, and genotoxicity. Recent advances and future perspectives.

7. Scientific Seminars
Critical analysis and discussion of scientific articles addressing recent applications of recombinant DNA technology and gene therapy. Preparation and presentation of student-led seminars.

• Giacca M. Terapia Genica. Springer, 2011.
• Kumar S. Textbook of Gene Therapy: Principles and Applications. Springer, 2026.
• Teaching materials provided by the instructor and selected scientific articles from the international literature.

Assessment consists of an oral examination and the preparation and presentation of a seminar based on a scientific article related to the course topics.

The oral examination aims to assess students’ knowledge and understanding of the course contents, their ability to establish connections between different aspects of recombinant DNA technology and gene therapy, and their use of appropriate scientific terminology.

The seminar aims to evaluate students’ ability to critically analyze scientific literature, summarize and present experimental findings, and discuss the potential and limitations of the approaches examined.

The final grade, expressed on a 30-point scale, will take into account the knowledge demonstrated during the oral examination, the quality of the seminar presentation, and the ability to critically analyze and discuss scientific findings.

1. What are the main steps of gene cloning, and what are the essential features of a cloning vector?
2. What are the main differences between cloning vectors and expression vectors?
3. Which elements should be considered when designing an expression cassette for eukaryotic cells?
4. What are the main differences between in vivo and ex vivo gene therapy?
5. Which strategies can be used to introduce a therapeutic gene or modulate its expression?