Learning objectives
KNOWLEDGE AND UNDERSTANDING ABILITY.
The course aims to implement the knowledge and skills related to the main genetic methods for the production of recombinant proteins, such
as drugs, vaccines and biomolecules of industrial interest, in prokaryotic
and eukaryotic microorganisms, in insect cells and in animal cells. During
the course the student will also learn the classical and molecular genetic
methodologies for the study of gene functions and interactions.
CAPACITY TO APPLY KNOWLEDGE AND UNDERSTANDING
At the end of the course the student should be able to design and deal with the recombinant production of proteins in the different expression systems. It should also be able to design simple experiments to study the
functions of essential and non-essential genes and their interactions in the yeast model system.
AUTONOMY OF JUDGMENT.
The student will be able to process what he has learned and recognize situations and problems in which the genetic methods described can be
used.
COMMUNICATION SKILLS
The student will acquire the ability to express himself with appropriate scientific language in the description of the methodologies treated and in
the discussion of experimental results. The student's ability to express himself in a clear and.
LEARNING ABILITY
The student will be able to deal independently with subsequent studies that require knowledge of genetic and molecular methodologies, and will
be able to apply the acquired study method to other areas.
Prerequisites
A basic knowledge of genetics, molecular biology and recombinant DNA technology is required.
Course unit content
The course aims to present the main methodologies and the most appropriate experimental approach for the large-scale production of recombinant proteins by exploiting different biological systems for the
expression of the cloned gene; from bacterial cells to animal cells. The main applications of biotechnological compounds such as hormones, cytokines, enzymes with therapeutic properties will be discussed. To address the recombinant production of vaccines and monoclonal antibodies, we will recall the fundamental notions of the mechanisms underlying the pathogenicity of bacteria and viruses and the genetic strategies that allow the diversification of immune responses to antigens.
The classical and molecular genetic methodologies for the study of gene functions and interactions in the "yeast model system" will also be presented.
Full programme
GENETIC ENGINEERING AND HETEROLOGOUS PROTEIN PRODUCTION
-Isolation of a gene of interest. Mutagenesis techniques.
THE CHOICE OF THE HOST:
-Analysis and optimization of the gene expression in prokaryotes. Gene
expression
from strong and regulatable promoters. Fusion proteins: cleavage and
use.
Increasing protein stability and secretion.
-Analysis and optimization of the gene expression in eukaryotic
microrganisms.
Yeast expression vectors, constitutive and regulatable promoters.
Secretion of
heterologous proteins in S. cerevisiae. Other yeast expression systems:
Pichia
pastoris and Kluyveromyces lactis.
-Baculovirus-insect cells expression system.
-Mammalian cell expression systems. Transient and stable expression.
Promoters
and reporter genes. Viral vectors.
THE PRODUCTS OBTAINED WITH THE RECOMBINANT DNA TECHNOLOGY
-Pharmaceuticals and enzymes.
-New vaccines.
-Monoclonal antibodies as therapeutic agents. Production of antibodies in
microrganisms.
STUDY OF GENE EXPRESSION AND FUNCTION.
-Northern Analysis, reporter genes, analysis using arrays.
-Gene interactions: suppressors, synthetic lethal mutations, mutations
gene dosage
dependent.
-The two-hybrid system
Bibliography
Glick, Pasternak and Patten “Molecular Biotechnology” ASM press IV edizione;
Dale and von Schantz "Dai geni ai genomi" Principi e applicazioni della
tecnologia del DNA ricombinante” Edises;
Watson J.D et al “DNA
ricombinante” Zanichelli. -Original papers provided by the teacher .
Teaching methods
The course includes lectures on specific topics of the program with the help of power point presentations. The teaching material used in the lessons and any other useful resources for the preparation will be provided to the student through the Elly platform. Course enrollment is required to access these online resources.
Assessment methods and criteria
The learning assessment will be done through a traditional oral exam which will consist of 3 questions. In the assignment of the final grade will be taken into account the level of knowledge of the different topics (insufficient, superficial, complete and in-depth), the student's ability to apply the theoretical concepts and formulate links between the various topics (sufficient, good, excellent) and of mastery of expression (deficient, simple, clear and correct).
Other information
- - -
