APPLIED BIOCHEMISTRY AND BIOCHEMICAL TECHNOLOGIES FOR BIOTECHNOLOGIES

Module TECNOLOGIE BIOCHIMICHE PER LE BIOTECNOLOGIE

The Molecular Diagnostic Technologies have broad application in the biomedical field and the course aims to give the student the knowledge of methods and tools to be used for the detection of gene mutations and genomic alterations necessary for diagnostic, prognostic and predictive of response before the start of targeted therapeutic protocols

Oral lectures integrated by laboratory activities. Should teaching be carried out in mixed mode or remotely, it may be necessary to introduce changes with respect to previous statements, in line with the programme planned and outlined in the syllabus. 

Knowledge of chemistry and biochemistry, cellular and molecular biology, genetics, anatomy and physiology acquired during the three-year degree

Weekly attendance in the first semester, first academic year as required by the Degree Course Regulation.

Biological samples: types, blood and biological samples, processing and storage. Extraction and purification of nucleic acids from human cells by chromatographic columns for gel filtration, ion exchange and affinity. Purification of nuclec acids with magnetic beads. Quantitative assay for nucleic acids using spectrophotometric and fluorimetric methods. Technologies based on the complementarity of the purine and pyrimidine nucleotides. Synthesis of cDNA by reverse transcription. Polymerase chain reaction (PCR). Qualitative PCR or End Point and quantitative PCR or Q-Real Time PCR. Programs for the design of primers to be used for PCR reactions. Evaluation of gene expression in real time by the method of 2-ΔΔCT. for Allelic discriminationusing the Real Time PCR. Restriction and Modification enzymes. Agarose gel electrophoresis for the separation of nucleic acids. Capillary electrophoresis for acid nucleic sequencing and analysis of microsatellite. Protein extraction and analysis. Electrophoresis of proteins. Western blot. DNA- and RNA-microarrays: Preparation of samples of DNA or RNA for the global analysis of the genome and transcriptome. Molecular analysis of the human karyotype microarray genomic photolithography-based probes to "Single Nucleotide Polymorphism-SNP" and "Copy Number Variation-CNV". Using algorithms and softwares for the interpretation of the data (CN state, Allele difference). Transcriptome analysis. Microarray expression to oligonucleotides obtained by photolithography synthesis. Array-CGH. Using algorithms and softwares for data interpretation (RMA and SAM). Application of the "microarray" technology for the study of solid and hematological tumors. Acid Nucleic First-generation sequencing technology, second generation and third generation. Methods for the preparation of libraries for the exome analysis and specific regions using a multiplex. Methods of clonal amplification. Techniques for the detection of incorporated nucleotides. Bioinformatics and statistical methods for interpreting data obtained from second-generation sequencing platform (NGS). Implementation of the first generation sequencing techniques for the detection of mutations with prognostic and predictive significance of response to therapy. Application of second generation technologies for the analysis of solid and hematological tumors. Detection of chromosomal aberrations by MLPA technology (Multiplex Ligation-dependent Probe Amplification). Human cell cultures.

1. Additional material supplied by the teacher; 2) Fundamental Molecular Biology; Lizabeth A. Allison, Editore: John Wiley & Sons Inc; 3) Online courses: http://www.dsf.unict.it/index.php?page=progetto-phar-in; "Riparazione del Dna e mutagenesi nel cancro" (D.F. Condorelli); Mismatch repair (MMR) instabilità dei microsatelliti (V. Barresi); "Tecnologie avanzate per la Bio-Medicina Genomica" (Vincenza Barresi). 

Oral exam. Learning assessment may also be carried out on line, should the conditions require it.

The evaluation of the knowledge acquired takes place through an oral exam. The grade is expressed on a scale of thirty, up to a maximum of 30/30 cum laude.

The following parameters will be taken into account to assign the final grade:

• Score 29-30 with honors: The student demonstrates in-depth knowledge of the topics, promptly and correctly integrates and critically analyzes the situations presented, independently solving even very complex problems. He has excellent communication skills and competently masters the language adopted in clinical biochemistry and clinical molecular biology.

• Score 26-28: The student has a good understanding of the topics, is able to integrate and analyze the situations presented critically and logically, explains the topics clearly using appropriate scientific language.

• Score 22-25: The student has a fair understanding of the topics, even if limited to the main areas. He is able to integrate and critically analyze the situations presented, although not always in a linear way, and presents the topics quite clearly with moderate linguistic competence.

• Score 18-21: The student has minimal knowledge of the topics, has a modest ability to integrate and critically analyze the situations presented and presents the topics clearly enough, although his/her linguistic competence may be poorly developed.

• Exam not passed: The student does not have the minimum required knowledge of the fundamental contents of the course. The ability to use a specific language is minimal or non-existent and the student is not able to independently apply the knowledge acquired.

Applications of first generation sequencing. Microarrays analysis applications. Parameters for the detection of CN-LOH. Application of "NGS" sequencing for RNA analysis.