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Practical 3 07

Practical 3 07






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    Practical 3 07 Practical 3 07 Presentation Transcript

    • Practical 3 Meiosis Gametogenesis
    • Chromosomal sex determination
      • Drosophila type
      • Male – XY
      • Female - XX
    • Sexual reproduction Life cycle: haploid phase (gametes) diploid phase (somatic cells) 2n 2n meiosis meiosis n n Gametes (haploid) sperm oocyte fertilization 2n n + n = 2n zygote ♂ ♀
    • Recommended website
      • http:// genetics.gsk.com/chromosomes.htm#chromo
    • Written test
      • 8 minutes
      • Don't forget to put down your name, your group and the test version.
      • In multiple choice questions more than 1 statement could be correct.
      • Don't write anything on the question sheet!
    • Meiotic division 2n 1 st meiotic division 2 nd meiotic division n n n n n n
    • First meiotic division
      • Heterotypic or reduction
      • Prophase
        • Leptotene
        • Zygotene
        • Pachytene
        • Diplotene
        • Diakinesis
      • Metaphase
      • Anaphase
      • Telophase
    • Leptotene
      • Beginning of prophase
      • Chromosomes can't be distinguished.
      • Condensation of chromosomes begins.
    • Zygotene
      • Attachment of homologous chromosomes – the chromosomes begin to synapse.
      • Assemblage of chromosomes is enabled by synaptonemal complex .
    • Pachytene
      • Bivalents or tetrades are present.
        • 1 bivalent = 2 chromosomes
        • 1 tetrade = 4 chromatids
        • bivalent = tetrade
      • Crossing-over between homologous chromosomes is possible.
    • Crossing-over
      • Reciprocal exchange of chromatid segments between chromosomal homologues
    • Importance of the crossing-over
      • New combinations of alleles of genes that are located on the same chromosome.
      • New allele combinations lead to higher variability of the offspring (important in evolution).
      • This increases the potential for genetic variability between members of the same species .
    • Chromosomes X and Y
      • are attached in pseudoautosomal regions (PAR) during the 1 st meiotic prophase
      X Y SRY – sex determining region – is located on chromosome Y in the neighborhood of PAR. X Y
    • Task A :
      • A n infertile man with azoospermia had been examined i n cytogenetic lab. He has no other striking phenotypic features.
      • The cytogenetic analysis revealed abnormal finding because his cells contained two X chromosomes while chromosome Y is missing.
      • Explain this surprising result.
    • Crossing-over in PAR
      • Nonreciprocal crossing-over can result in translocation of the SRY gene from the chromosome Y to chromosome X
    • Task A – result male female Normal combination of gonosomes X;Y translocation XX XY „ XX-male“ „ XY-female“
    • Hermaphroditism – possible consequence of sex chromosome abnormalities
    • Prophase of the 1 st meiotic division next periods
    • Pachytene
    • Diplotene
      • Condensation of chromosomes continues
      • Homologous chromosomes are divided
      • Disassembling of synaptonemal complex
      • At the end of diplotene homologues are connected only in crossover sites. These sites are called chiasmata (singular: chiasma).
    • Diakinesis
      • Final period of the 1st meiotic prophase.
      • Chiasmata are transferred to terminal chromosomal regions.
      • Under specific staining chromosomes could be well distinguished under the microscope.
      • Nuclear membrane disappears.
    • Terminal phases of meiotic division prophase metaphase + anaphase 2 nd meiotic division (homeotypic) 1 st meiotic division (heterotypic) Separation of double-chromatid homologous chromosomes
    • Metaphase I Metaphase I
    • Anaphase I X Anaphase II
    • Gametogenesis
      • Spermatogenesis – sperm development
      • Oogenesis – oocyte development
    • Spermatogenesis n n 1st meiotic division 2nd meiotic division n n primary spermatocyte secondary spermatocyte sperm n n 2n
    • Oogenesis n n 1st meiotic division 2nd meiotic division n n primary oocyte secondary oocyte mature oocyte n n 2n polar bodies polar body
    • A schedule of oocyte development Prophase I fetus childhood – dictyotene mature ovary fertilization
    • Spermatogenesis X Oogenesis
    • Task B : Risk of chromosomal nondisjunction defects
      • Compare possible risk of chromosomal abnormalities for two couples that visited the genetic counselling clinic:
        • Couple 1: female (21 years), male (38 years)
        • Couple 2: female (36 years), male (32 years)
      • Which family has higher risk of numerical chromosomal aberrations?
    • Task B – result
      • Oogenesis lasts much longer than spermatogenesis – risk of chromosomal nondisjunction defects is age-related.
      • The second couple has higher risk due to 36 years old female.
      • In pregnant females older than 35 years prenatal examination of the fetus is recommended.
      35 years … risk 1/365 Age-related risk of the Down syndrome child Mother´s age
    • Practical tasks
    • Task 1 : Observation of meiosis in the grasshopper testis
      • The slide was prepared from the testis of the grasshopper. After removing from the individual the tissue was fixed with methanol + glacial acetic acid (3 : 1) and stained with the Giemsa-Romanowski solution. After dehydration the tissue was mounted in the Canada balsam.
      • Find the spermatocytes on the slide using 10x objective lens.
      • Change the objective magnification into 40 or 45x and observe different phases of meiosis. For their determination use adjacent photos.
    • Task 2 : Disjunction of the sex chromosomes during meiosis
      • Redraw the scheme of disjunction of the sex chromosomes during meiosis of the male to your materials and fill it.
      • In the first scheme (A) distinguish the chromosomes according to their shape and number of chromatids.
      • In the second scheme (B) designate the chromosomes using letters X or Y no matter of the number of chromatids.
      A B XY
    • Task 3 : Changes of nuclear DNA concentration during the cell cycle
      • Contrast the two types of cell division (mitosis X meiosis) by completing the following graphs below.
      • Redraw the picture to your materials and fill curves demonstrating changes in the DNA content per one cell:
        • during the cell cycle followed with mitosis
        • during the cell cycle followed with meiosis.
      • (Value „C“ on y-axis represents DNA content per one cell in G1 phase.)
    • Task 3 : Changes of nuclear DNA concentration during the cell cycle G 1 G 2 S M G 1 C 2C G 1 G 2 S M 1 M 2 G 1 Cell cycle + mitosis Cell cycle + meiosis Nuclear DNA content
    • Results
    • Task 3: Observation of meiosis in the grasshopper testis
    • Task 4: Disjunction of the sex chromosomes during meiosis Scheme A Scheme B XY X Y X X Y Y
    • Task 5: Changes of nuclear DNA concentration during the cell cycle G 1 G 2 S M G 1 C 2C G 1 G 2 S M 1 M 2 G 1 Cell cycle + mitosis Cell cycle + meiosis Nuclear DNA
    • Assisted reproduction
      • A group of methods that allows reproduction of infertile couple that can't be therapeuted by common medical treatment (e.g. using drugs of by psychological intervention)
      • The assisted reproduction could be also used for prevention of genetic defects in families with normal fertility but with increased risk of genetically conditioned disorder for the offspring.
    • Assisted reproduction methods
      • Intrauterine artificial insemination
        • application of sperm to the uterus of the female.
      • In vitro fertilization and embryo transfer
        • fertilization is performed outside the organism of the mother
    • Artificial insemination
      • homologous – the female is fertilized with sperm of her husband;
      • sperm of anonymous donor are applied – the donor and the couple remain unknown to each other
    • In vitro fertilization
      • Application of gonadotropins (= fertility drugs) induce maturation of numerous oocytes.
      • The oocytes are removed from ovaries by biopsy. In a special dish they are mixed with sperms of the husband.
      • After short term cultivation in medium with nutrients 3 embryos are transferred to the female's uterus.
    • ICSI (intracytoplasmatic sperm injection)
    • ICSI protocol
    • Preimplantation diagnostics
      • Genetic or other examinations of oocytes or embryo before transfer to the mother's
      • We use
        • Polar body
        • 1 – 2 cells from 8-cell embryo
        • More cells from blastocyst
    • Presentation Ethical issues of assisted reproduction
    • Next seminar
      • In vitro cell cultivation
      • Recommended websites:
        • http://www.research.umbc.edu/~jwolf/method5.htm
        • http://homepages.gac.edu/~cellab/chpts/chpt12/intro12.html
      • Be careful when handling cell cultures!
      • Test: previous topics (only seminars)
    • See you next week!