Teaching code

4S003719

Credits

12

Coordinator

Alessandra Maria Bossi

Language

Italian

Moodle Seminars 0

The teaching is organized as follows:

Learning outcomes

Transcriptomic (theory part)
The theory part of this course has as aim to explain what a transcriptomic analyses is and for which purpose it can be applied. Furthermore it aims to provide knowledge about the different approaches and technique which can be applied for a transcriptomic analysis.

Transcriptomic (laboratory part) The laboratory part of this Course has as main aim to provide to students the knowledge for the cor-rect design, application and interpretation of a transcriptomic analysis. Students will have the oppor-tunity to approach and discuss the experimental design for a transcriptomic experiment, to deal with preparation of samples for RNASeq analysis and will apply bioinformatics tools to interpret tran-scriptomic data.

Metabolomics (theory part)
- define the “targeted” and “untargeted” metabolomics approaches;
- discriminate the critical points of an experimental design;
- organize a plan for the extraction and analysis of metabolites suitable for the metabolites/class of metabolites of interest;
- describe the key elements for the raw data process till the production of the FQM (Feature Quantifi-cation Matrix);
- use statistical approaches suitable to the experimental problem;
- apply the metabolomics analysis to the investigation of the biological complexity of cells/ tissues/ or-gans/organisms, from the point of view of the small molecules, as intermediates or end
products of the metabolism;
- use an appropriate scientific language.

Metabolomics (laboratory part)
- Simulation of a “real world” experimental design of a metabolomics and transcriptomics experiment: the students, organized in work group, will receive an experimental problem and virtual resources (biological resources, department facilities, lab instrumentation and budget), and will be required to lay down an experimental plan;
- extraction of plant medium and low polar metabolites, cleaning of samples through Solid Phase Ex-traction, visit to the LC-MS facilities;
- LC-MS-based -untargeted metabolomics: recognition of molecular ions in LC-MS chromatogram; fragmentation trees; metabolite identification through m/z values and fragmentation trees;
- chromatogram processing and Feature Quantification Matrix building;
- multivariate and univariate data analysis.

Proteomics (theory part)
The unit is intended to provide the theoretical bases to tackle the proteomic analysis. Starting from the description of the physico-chemical characteristics of the sample and of the related methods to fractionate the sample, the course develops later in the account of the principal and most exploited separative methods serving the proteomic analysis (gel, gel free), in the elucidation of mass spec-trometric analysis for the proteome and of the in silico methods for identification of the proteins. Fur-thermore, the main methods used for quantitative proteomic analysis will be examined.

Proteomics (laboratory part)
Provides proteomic analysis experimental protocols, addressing the steps needed to map the prote-ome. The laboratory is divided into the following phases: experimental design, sample processing, separation methods in gel, protein identification by means of analysis in silico.

Program

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MM: MODULO II
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------------------------ MM: Metabolomica laboratorio ------------------------ -Simulation of a “real world” experimental design of a metabolomics and transcriptomics experiment: the students, organized in work group, will receive an experimental problem and virtual resources (biological resources, department facilities, lab instrumentation and budget), and will be required to lay down an experimental plan; -extraction of plant medium and low polar metabolites, cleaning of samples through Solid Phase Extraction, visit to the LC-MS facilities; -LC-MS-based -untargeted metabolomics: recognition of molecular ions in LC-MS chromatogram; fragmentation trees; metabolite identification through m/z values and fragmentation trees; -chromatogram processing and Feature Quantification Matrix building; -multivariate and univariate data analysis. ------------------------ MM: Proteomica laboratorio ------------------------ Sample preparation. Separation by means of Gel-Based Proteomics. Proteins and peptides mass spettrometry and mass/mass spectrometry. Protein identification methods and Protein Database Search. ------------------------ MM: Proteomica teoria ------------------------ The preliminary requisite to study proteomics, is to possess basic concepts of biochemistry and of the principal methods utilized in biochemistry. 1. Introduction. 2. Sample preparation. 3. Gel-Based Proteomics. 4. Gel-Free Proteomics. 5. Saple pre-fractionations and analysis of low abundant proteins. 6. Proteins and peptides mass spettrometry and mass/mass spectrometry. 7. Protein identification methods and Protein Database Search. 8. Quantitative Proteomics. 9. Examples of proteomics applied to food.
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MM: MODULO I
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------------------------ MM: Metabolomica teoria ------------------------ -the experimental design; -metabolite extraction and preparation for LC-MS; -data formats and conversion; -chromatogram analysis and metabolite annotation; -data preprocessing; the effect of the technique of data normalization; -data processing (with MZmine): m/z feature extraction, chromatogram deconvolution and alignement; production of the Feature Quantification Matrix; -data analysis through multivariate analysis (PCA, PLS-DA, OPLS-DA) and univariate analysis. ------------------------ MM: Trascrittomica laboratorio ------------------------ ------------------------ MM: Trascrittomica teoria ------------------------

Examination Methods

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MM: MODULO II
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------------------------ MM: Metabolomica laboratorio ------------------------ After a public presentation and a critical discussion of the work of each groups, each student will have a score (from 0 to 2) that will additively contribute to the final partial score of the metabolomics examination. ------------------------ MM: Proteomica laboratorio ------------------------ The exam is aimed at verifying the acquisition by the student of the skills described in the objectives (knowledge of the protein material and of the sample under analysis, mastery of the methods to extract proteins from the sample, decision-making skills in the choice of the separative method; analysis in mass spectrometry and spectra reading, in-silico database research skills for protein identification, result validation skills, ability to define a quantitative strategy of proteomic analysis). The exam is written, open questions, 2 hours. The written exam is integrated, upon student’s request, with an oral verification. The exam will be evaluated on the basis of the student's ability to answer the questions correlating concepts and methods in an appropriate manner and, where proposed, detailing the meaning of equations, or reporting relevant graphs. ------------------------ MM: Proteomica teoria ------------------------ The exam is aimed at verifying the acquisition by the student of the skills described in the objectives (knowledge of the protein material and of the sample under analysis, mastery of the methods to extract proteins from the sample, decision-making skills in the choice of the separative method; analysis in mass spectrometry and spectra reading, in-silico database research skills for protein identification, result validation skills, ability to define a quantitative strategy of proteomic analysis). The exam is written, open questions, 2 hours. The written exam is integrated, upon student’s request, with an oral verification. The exam will be evaluated on the basis of the student's ability to answer the questions correlating concepts and methods in an appropriate manner and, where proposed, detailing the meaning of equations, or reporting relevant graphs.
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MM: MODULO I
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------------------------ MM: Metabolomica teoria ------------------------ The examination will be an oral test aimed to verify : -the overall degree of knowledge of the subject of this course; -the scientific language skills. ------------------------ MM: Trascrittomica laboratorio ------------------------ The exam will be oral aiming to ascertain the students’ knowledge on the topics of lectures and lab practices. One third of the colloquium will be dedicated to test knowledge on topics treated in the lab part of the course. Student will discuss experimental planning or applications of transcriptomic analysis or apply tools for transcriptomic data analysis faced during the lab part. The exam will be same for both attending and non-attending students. ------------------------ MM: Trascrittomica teoria ------------------------ The exam will be oral aiming to ascertain the students’ knowledge on the topics of lectures and lab practices. Two third of the colloquium will be dedicated to test knowledge on topics treated in the theory part of the course. The exam will be same for both attending and non-attending students.