Syllabus
* 1st day - introduction, methods of study of DNA, differences among molecular markers - DNA extraction from plant material (CTAB method, commercial kit) - fresh and dried material - electrophoresis, making agarose minigels, use of DNA ladders - working with documentation system Kodak Gel Logic 100 - determining of DNA concentration using UV spectrophotometer (Nanodrop), DNA dilution * 2nd day - PCR amplification, making PCR premix, methods of PCR optimization - working with diverse termocyclers - making new PCR programs, use of gradient block for PCR optimization - PCR method - optimization using gradient block - electrophoresis of PCR bands and their visualization - working with the software for gel analysis (KODAK 1D Image Analysis Software) - estimation of the lengths of PCR bands * 3rd day (PCR-RFLP) - PCR amplification using optimal annealing temperature- restriction of PCR bands with restriction endonucleases
* 4th day - dominant molecular markers and data evaluation - theory- analysis of test gels, preparing binary data matrix- evaluation of the data - FAMD software- diversity calculation, PCoA, trees, networks, AMOVA - FAMD, AFLPdat, SplitsTree * 5th day - continue with data analysis- Bayesian model-based approach - STRUCTURE software- visualization of the STRUCTURE results - Structure-sum, Distruct
Annotation
The lectures/practicals are given in English if non-Czech speaking students are enrolled. A practical introduction to the methods of molecular markers in DNA lab at the Department of Botany.
Students will learn methods of DNA extraction from the plant material and PCR optimization. In the second part PCR-RFLP method on chloroplast DNA is demonstrated.