1. komponenta

Lecture type	Total
Lectures	30
Practicum	45
Seminar	30

* Load is given in academic hour (1 academic hour = 45 minutes)

Course Content:

1. Introduction to the basics of laboratory work, the laboratory safety level and model organisms
Behavior in the laboratory, categorization into four safety levels depending on model and transgenic organisms. Methods of sterilization, disposal and personal protection.
2. Cell culture basics and reporter genes
The types of cells, growing conditions, methods of transgene introduction, expression systems, vectors, types of reporter genes and methods of their application and detection, flow cytometry.
3. Methods for protein research
Isolation of proteins, methods for the determination of protein concentration, SDS PAGE electrophoresis, Western blotting and detection of the specific protein. Methods for determining protein-protein interactions by immunoprecipitation, FRET techniques, using yeast two-hybrid assay. Fundamentals and applications of immunocytochemistry and immunohistochemistry.
4. DNA hybridization methods
Basics of fluorescence in situ hybridization and similar methods, applications in diagnostics.
5. Real time PCR
The basics of the method, the type and design of primers, analysis of results, the use of qPCR in the diagnosis and monitoring of HIV infection and Bcr-Abl chimeric transcript in leukemia patients.
6. Sequencing
Sanger sequencing method, the second-generation sequencing (pirosequencing) and third generation sequencing (nanopore). Advantages and disadvantages of different methods and historical overview.
7. Analysis of polymorphisms in genomic and mtDNA
VNTR, STR and SNP polymorphisms and their analysis in anthropology and forensics. Structure and analysis of hypervariable region of mitochondrial DNA and applications - paternity testing, the origin of the human populations, and criminal investigations.


Lab work:

Each student participates independently in the 4 exercises.
1. Transfection of human cells by GFP vector - exercise involves working with cell culture, feeding and passaging cells, transfection using calcium phosphate and analysis by fluorescent microscopy, photography and image processing with Photoshop
2. Protein analysis - a student can choose between Western analysis, immunoprecipitation and immunocytochemistry.
3. FISH analysis of DNA
4. Application of the methods - the student chooses one of the exercises offered by the collaborating institutions. They offer the following exercises: flow cytometry and qPCR in the diagnosis of infectious diseases (Hospital for Infectious Diseases Fran Mihaljevic), qPCR in the diagnosis of leukemia (Dubrava), STR analysis in paternity testing (Genos), analysis of mtDNA (Institute of Anthropology), and DNA sequencing (Rudjer Boskovic).


Seminars:

1. writing and presenting the research proposal
2. presentations of lab work methods and results

1. Ambriović Ristov A. Metode u molekularnoj biologiji, Institut Ruđer Bošković 2007.
   Keith Wilson i John Walker: Principles and Techniques of Biochemistry and Molecular Biology, Cambridge University Press, 2011.
   Kathy Barker: At the bench: A laboratory navigator, Cold Spring Harbor Laboratory Press 2005.

Mandatory course - Regular study - Molecular Biology