BIOCHEMISTRY AND APPLIED BIOCHEMISTRY F3150  Farmacia

Part II : Applied Biochemistry  CFU 5 

Scopes and objectives: the course covers the fundamental molecular mechanisms of the conversion of information contained in DNA to cellular macromolecules (DNA, RNA, protein biosynthesis). The topics include the recombinant DNA technology and the basic principles of the methods  used in studying proteins, enzymes, nucleic acids and clinical biochemistry.

The exam is oral. Final mark is the result of the marks of part I and part II.

Requirement: basic knowledge of organic chemistry

 

PROGRAM:

 

BASIC MOLECULAR BIOLOGY:

- DNA structure, DNA A, B, Z, H; topological properties and supercoiling; the structure of chromosome in prokaryotic and eukaryotic cells; Telomeres and telomerase.

- DNA replication in prokaryotes and eukaryotes; reverse transcription; recombination and DNA repair

- transcription: synthesis of the different types of RNA and processing of mRNA (caps, poly-A tail, splicing, alternative splicing); Si RNA; factors affecting transcription

- translation:  genetic code, tRNA, aminoacyl tRNA tranferases and protein biosynthesis.

RECOMBINANT DNA TECHNOLOGY

- restriction enzymes, plasmids, DNA cloning, cDNA and genomic libraries

MOLECULAR DIAGNOSIS

- molecular probes, DNA fingerprints, Southern blotting, Northern blotting.

- DNA sequencing

- PCR.

ELECTROPHORESIS of DNA and PROTEINS

- Basic principles

- electrophoresis of nucleic acid

- electrophoresis of proteins: SDS-PAGE, isoelectric focusing, two-dimensional electrophoresis; composition, functions and electrophoresis separation of serum proteins.

CHROMATOGRAPHY an d PROTEIN PURIFICATION

- gel chromatography, ion exchange and affinity chromatography.

SPECTROPHOTOMETER METHODS FOR PROTEIN, ENZYMES and NUCLEIC ACID STUDIES

- general principles and definitions

- examples of absorption spectra of biological molecules: proteins, nucleic acids, NAD, hemoproteins

- protein and DNA assays

- enzyme assays: rate of enzyme-catalyzed reactions and enzyme concentration, units of enzyme activity, specific activity. Examples of enzyme assays: cytochrome c reductase, malate dehydrogenase, maltase, proteases, aminotransferases. Enzymes as reagents: enzymatic assay of cholesterol and glucose.

 

IMMUNOLOGICAL ASSAYS

- general concepts, immunoprecipitation and immunolabelling

-RIA and ELISA

- western blot

 

 

RESULTS EXPECTED

 Learning of the structure-function relationship of biological macromolecules and of the biochemical flux of information. Learning of the basic methods used in the biochemical studies and in the recombinant DNA technology

 

Course books:

K.WIlson, J. Walker: Biochimica e biologia molecolare-principi e tecniche.- Raffaello Cortina editore.
C.K. Mathews et al, Biochimica, Casa Editrice Ambrosiana
Campbell, Farrel, Biochimica, EdiSES