66435 - Applied Molecular Biology

Learning outcomes

The students will be able to operate by ourself in a clinical-pathology laboratory. Furthermore, they will be able to effectively join a lab team, to organize the diagnostic procedures and to perform clinical and molecular assays.

Course contents

To learn basic protocols of proteomics (extraction, quantification separation of sub-proteomes, 1D and 2Delectrophoresis, western blotting, mass protein identification) and metallomics (metals and metalloproteins identification); to apply these protocols in the diagnostic field.

To learn basic techniques of molecular biology (DNA isolation, Polymerase Chain Reaction, Sanger sequencing, analysis of fragments, single nucleotide polymorphisms (SNPs). To learn the fundamentals of the main statistic computations (variability and diversity indices, phylogenetic trees, multivariate and Bayesian analysis). Acquired knowledge will be used to assess conservation projects related to wildlife and forensic investigation.

To learn methods for performing molecolar assay aimed at diagnosing neoplastic and genetic diseases as well as genotyping microsatellites for parentage verification in domestic animals. To learn skills for performing automated and manual clinico-pathological analysis and in particular clinical-chemistry, hematology, electrophoresis, urinalysis, fecal examination, hematological and cytological staining, coagulation assay. To learn how to organize and manage a clinical laboratory.

Molecular diagnosis. 1.Genetics of immunoglobulins; definition of lymphoid marker and Ig and TCR gene rearrangements. Heteroduplex analysis and fragment analysis as diagnostic method for detecting Ig and TCR clonal rearrangements. Methods for purifying DNA for the molecular diagnosis. 2.Role of somatic mutations in cancerogenesis. “Drivers” and “passengers” mutations. Techniques used for the highly sensitive detections of somatic mutations in neoplastic tissues. 3.Repeated DNA sequences. Microsatellites as polymorphic markers for parentage testing and pedigree verification. 4. Techniques for genotyping single nucleotide variations: restriction enzymes, real-time PCR, High resolution melting curve analysis, allele-specific primer extension methods.

Readings/Bibliography

Powerpoint lectures will introduce the practical sessions. For some cases, brief videos will be used to illustrate the techniques.

Teaching methods

For practical work, groups composed by 2 or 3 students will perform the molecular assays.

Assessment methods

Individual skills will be monitored during practical activity.

The final grade will be composed of the active participation during lecture and laboratory activities and a written exam about lecture topics.

This teaching is part of an Integrated Course, the final grade will be proportional to the CFU of each teaching

35106 Policy and Legislation in Animal Biotechnology 3CFU 20%

66437 Applied Experimental Biology 6CFU 40%

66435 Applied Molecular Biology 6CFU 40%

Teaching tools

The course is strictly practical. Practical sessions will be introduced by a short lecture to fournish the background of the aims of the assay. Students will learn the extraction methods to purify nucleic acids from clinical samples and the methods to amplify the target of neoplastic diseases and to perform allelic discrimination assays. Also the amplification of polymorphic traits as microsatellites will be afforded during the course.

The organization and managment of a clinical-pathology Laboratory as well as le operative instructions on how to perform clinico-pathological assays will be presented at the Veterinary Clinical-pathology laboratory. The students will operate ourselves.

Office hours

See the website of Gloria Isani

See the website of Fabio Gentilini

See the website of Giulia Andreani