Syllabus

1) Current views on the chromatin structure and its topology in the nucleus

- topologically associated domains (TADs), lamin associated domains (LADs), chromosome territories, functional significance

2) Fluorescence in situ hybridization – the principles and the protocol overview

- direct and indirect labeling of FISH probes and probe detection (advantages and limitations)

- principles of probe labeling (nick translation, random priming, DOP PCR, primed in situ labelling (PRINS) etc.)

- effect of formamide on DNA denaturation, the role of other chemical agens in the basic FISH protocol

- the resolution of FISH-based techniques, stringency

3) Current markers and methods of molecular cytogenetics

- tandemly-repeated sequences, satellite DNA (satDNA), Cot DNA, microsatellites, transposable elements (TEs), telomeres and interstitial telomeric sites (ITSs), multigene families (histone genes, snDNA, rDNA) and their bearing to karyotype analysis

- mechanisms and models describing evolutionary dynamics of tandem repeats (concerted evolution, birth-and-death evolution, impact of gene conversion and unequal crossing-over along with recent insights into mechanisms of these phenomena)

- detailed physical mapping of adjacent or co-localized markers (fibre-FISH)

- mapping of single-copy regions – tyramide signal amplification FISH (TSA FISH), BAC (bacterial artificial chromosome) FISH

- oligo-paint FISH

- whole chromosome painting probes (WCP) and their intra- and interspecific (Zoo-FISH) application

- FISH with probes derived from whole genomic DNA- genomic in situ hybridization (GISH) and comparative genomic hybridization (CGH)

- utilization of next generation sequencing (NGS), available genomic data and particular databases for generation of new cytogenetic markers

- examples of integrated approach combining cytogenetic methods with data of other (phylogenetic, taxonomic, ecological, evolutional, genomic) approaches

4) Application of molecular cytogenetics in analyses of specialized chromosomes

- methods suitable for deciphering the emergence and evolution of sex/neo-sex chromosomes and B chromosomes, with emphasis on specific cases (Rumex, Drosophila, Orthoptera, Lepidoptera, spiders, fishes, frogs, reptiles, marsupials)

- unusual and extreme cases of sex chromosome differentiation

- methods suitable for unmasking master sex determining genes (or other sex-linked genetic content) including state-of-the-art sequencing and bioinformatic tools

- turnover of sex chromosomes and its high rate in cold-blooded vertebrates

5) Contribution of molecular cytogenetics to research of polyploidy, hybridization, asexual/unisexual reproduction and genome elimination

- applications and resolution power for resolving between auto- vs. allopolyploidy scenario in recent and older evolutionary events; evolutionary dynamics of post-polyploid genome restructuring, re-diploidization

- homoploid vs. allopolyploid hybrids, unique case of recurrent hybridization in sturgeons

- gynogenesis, hybridogenesis (with special emphasis on fishes and amphibians - Ambystoma, Cobitis, Pelophylax)

- genome elimination in frogs, fishes and nematods and associated battery of suitable methods for its investigation

6) Molecular cytogenetics and karyotype evolution

- mechanisms of karyotype evolution (chromosome rearrangements and their impact on reproductive isolation, karyotype orthoselection, meiotic/centromeric drive)

- trends of karyotype evolution in particular taxonomic groups (with emphasis on typical as well as outstanding examples)

- centromere repositioning

- microchromosomes

- polymorphisms, karyomorphs, chromosome races, cryptic species

- hybrid zones, introgression

- karyotype stasis

- unique patterns of karyotype evolution in polyploids and taxa with holocentric chromosomes

- mapping of conserved synteny and its bearing to reconstruction of ancestral karyotypes

- chromothripsis as a potential driving force behind karyotype evolution