Bi7200 Advanced methods in plant cytogenomics

Faculty of Science
Autumn 2025
Extent and Intensity
0/2. 2 credit(s) (plus extra credits for completion). Type of Completion: z (credit).
In-person direct teaching
Teacher(s)
RNDr. Terezie Malík Mandáková, Ph.D. (lecturer)
Guaranteed by
RNDr. Terezie Malík Mandáková, Ph.D.
Department of Experimental Biology – Biology Section – Faculty of Science
Contact Person: RNDr. Terezie Malík Mandáková, Ph.D.
Supplier department: Department of Experimental Biology – Biology Section – Faculty of Science
Prerequisites (in Czech)
Bi6270 Cytogenetics
Course Enrolment Limitations
The course is only offered to the students of the study fields the course is directly associated with.

The capacity limit for the course is 32 student(s).
Current registration and enrolment status: enrolled: 15/32, only registered: 0/32
fields of study / plans the course is directly associated with
Course objectives
This course develops key skills essential for modern research in plant genomics. Students will learn to visualize chromosomal structures in various developmental and cellular contexts and to integrate these observations with molecular and evolutionary data. They will gain a deeper understanding of how plant genomes change over time, how chromosomal rearrangements arise, and how these changes influence plant diversity and adaptation. The practical focus of the course encourages independent thinking, critical evaluation of results, and a creative approach to solving biological questions. The acquired skills are applicable in both academic and applied research, particularly in the fields of plant genomics, phylogenomics, breeding, and conservation of genetic resources.
Learning outcomes
Upon completion of the course, students will gain hands-on experience with advanced methods for visualizing and analyzing plant chromosomes. They will be able to prepare high-quality preparations of mitotic and meiotic chromosomes as well as interphase nuclei from various plant tissues. Students will learn to design and carry out experiments using FISH, GISH, and chromosome painting with different types of probes (e.g., repetitive DNA, rDNA, BAC clones, genomic DNA). They will master the use of fluorescence microscopy, digital documentation, and basic image data analysis. Students will understand the principles of hybridization and be able to interpret results in the context of plant genome structure, evolution, and diversity. The course will prepare them for independent laboratory work and active participation in research projects in plant genetics, cytogenomics, and evolutionary biology.
Syllabus
Day 1 – Introduction and sample preparation The first day introduces plant cytogenetics, highlighting differences between model and non-model species and the specific features of plant cells, such as the absence of centrioles, rigid cell walls, and the need for cell cycle synchronization. Students will learn about different types of chromosome preparations—mitotic chromosomes from root tips, meiotic chromosomes from anthers, and interphase nuclei from leaves—and how to plan experiments based on research questions, species selection, tissue type, probe type, and labeling strategy. The practical part includes collecting plant material at the correct developmental stage, fixation in Carnoy’s solution, enzymatic digestion using pectolytic enzymes, and slide preparation. Simultaneously, DNA is isolated from BAC clones or genomic sources, quality is checked, and probe labeling is initiated using nick translation with haptens or fluorochromes. Day 2 – Hybridization setup and principles This day focuses on preparing for hybridization and understanding the principles of in situ hybridization techniques. Students will learn the applications of FISH (for locating specific sequences), GISH (for distinguishing parental genomes), and chromosome painting (for studying structural details and evolutionary changes). Challenges such as high repetitive DNA content and chloroplast autofluorescence in plant genomes are discussed. Probe labeling is completed, fragmentation is checked via electrophoresis, and hybridization mixtures are prepared, potentially combining repetitive sequences, rDNA, genomic DNA, and BAC clones. DNA denaturation on slides and in probes follows, along with application of the hybridization mix, coverslip mounting, and overnight incubation at a stable temperature. Day 3 – Post-hybridization and interpretation The third day is dedicated to post-hybridization steps and interpreting results in the context of plant genome evolution. After hybridization, slides are washed in formamide and SSC solutions to remove non-specifically bound probes. Immunodetection is performed using labeled antibodies, such as avidin-Texas Red for biotin or anti-digoxigenin-Alexa488. Theoretical discussions cover the use of chromosome painting and its role in identifying chromosomal rearrangements such as fusions, inversions, and translocations. Students discuss homeologous relationships between species, ancestral karyotype reconstruction, and prepare for microscopy by analyzing expected signal patterns and their interpretations. Day 4 – Microscopy, image analysis, and presentation The final day focuses on microscopy, digital image analysis, and presentation of results. Students use fluorescence microscopes, select appropriate filters for each fluorochrome, adjust exposure settings, and capture images of hybridization signals. They document the localization of 45S rDNA, telomeres, parental genome differentiation via GISH, and BAC probe colocalization in CCP. Image data are analyzed using software such as ImageJ or Photoshop, including channel overlays, signal annotation, and distance measurements. Students prepare a presentation or poster summarizing their experiment, methodology, results, and interpretations. The course concludes with presentations, discussion, feedback, and suggestions for applying the methods in their own research.
Literature
    required literature
  • SCHWARZACHER, T. and HESLOP-HARRISON. Practical in situ hybridization. Bios, 2000. info
    recommended literature
  • Heitkam, T., & Garcia, S. (Eds.). (2023). Plant cytogenetics and cytogenomics: Methods and protocols. Methods in Molecular Biology, Vol. 2672. Springer. https://doi.org/10.1007/978-1-0716-3155-4
  • MANDÁKOVÁ, Terezie and Martin LYSÁK. Chromosome Preparation for Cytogenetic Analyses in Arabidopsis. Current protocols in plant biology. John Wiley & Sons, 2016, vol. 1, No 1, p. 43-51. ISSN 2379-8068. Available from: https://doi.org/10.1002/cppb.20009. URL info
  • MANDÁKOVÁ, Terezie and Martin LYSÁK. Painting of Arabidopsis Chromosomes with Chromosome-Specific BAC Clones. Current protocols in plant biology. John Wiley & Sons, 2016, vol. 4, No 2, p. 359-371. ISSN 2379-8068. Available from: https://doi.org/10.1002/cppb.20022. URL info
Teaching methods
The course is delivered as an intensive five-day laboratory block combining hands-on exercises, theoretical lectures, and independent student work. Emphasis is placed on active participation, with students mastering all steps—from chromosome preparation and hybridization techniques to microscopy and digital data analysis. Students work with modern laboratory equipment, including fluorescence microscopes and image analysis software. The course also includes work on individual projects or model tasks, allowing students to apply their skills in the context of their own research interests. The course concludes with a presentation of results, where students summarize their experimental procedures, interpret data, and discuss potential applications in their own research.
Assessment methods
The course is completed with a credit (pass/fail), awarded based on active participation, successful completion of practical tasks, and presentation of the student’s experimental results.
Language of instruction
Czech
Follow-Up Courses
Further comments (probably available only in Czech)
The course is taught annually.
The course is taught in blocks.
Teacher's information
The course is delivered as an intensive five-day laboratory block combining hands-on exercises, theoretical lectures, and independent student work.
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