Cytogenetics and Chromatin Structure

Cytogenetics

• Cytogenetics is the branch of genetics studying chromosome structure, function, and pathology.
• Chromosomes are structures in cell nuclei comprised of DNA and histones.

DNA Organization in the Nucleus

• Almost all DNA in eukaryotes is concentrated in the nucleus.
• Chromatin is a complex dynamic structure formed by DNA and proteins.
• Eukaryotic chromatin consists of DNA, histone proteins, non-histone proteins, and RNA (<10% of DNA mass).

Chromatin

• Euchromatin and heterochromatin differ in structure, DNA sequences, and activity.
  • Euchromatin is less condensed, open, accessible, with coding genes and is transcriptionally active.
  • Heterochromatin is condensed, closed, inaccessible, with repeating elements and is transcriptionally inactive.

Nucleosome Structure

• Nucleosomes are the basic structural and regulatory units of chromatin.
• A nucleosome is formed by DNA revolving around histone proteins.
• One nucleosome consists of:
  • A histone octamer (two molecules of four different histones: H2A, H2B, H3, and H4).
  • The histone octamer binds to approximately 200 nucleotide pairs of DNA.
  • One linker histone (e.g., H1) molecule is bound to one nucleosome in chromatin.
• Nucleosomes represent the first level of chromatin organization.

Levels of Chromatin Organization

• Chromatin undergoes various structural levels of organization, from the 10 nm chromatin fiber to condensed chromosomes:
  • 10 nm chromatin fiber: DNA wrapped around histone octamers.
  • 30 nm chromatin fiber: Nucleosomes packed together.
  • Chromatin loops: 30 nm fibers folded into loops.
  • Condensed chromatin: Tightly packed chromatin, forming chromosomes.

Chromosome Structure

• Chromosomes are condensed chromatin structures, visible during cell division.
• Centromere: Constricted region dividing the chromosome into arms (short (p) and long (q)).
• Telomeres: Protective caps at the ends of chromosomes.

Chromosome Types

• Chromosomes are categorized based on centromere location.
  • Metacentric: Centromere in the middle, resulting in equal-sized arms.
  • Submetacentric: Centromere slightly off-center, creating unequal arms.
  • Acrocentric: Centromere near one end, resulting in a very short arm.
  • Telocentric: Centromere at the end, with only one arm.

Chromosomal Numbering System

• Chromosomes are numbered based on their size and centromere location.
• Banding patterns are used to identify specific regions and sub-regions within a chromosome.

Karyotype

• Karyotype: An organized set of chromosomes arranged by size and shape.
• Each chromosome pair is marked with a number, except for sex chromosomes.

Chromosome Ideograms

• Ideogram: A graphic representation of a chromosome showing the positions of structures (centromeres, telomeres, shoulders) and genes.
• Banding patterns reveal the exact location of genes or chromosomal abnormalities.

Use of Cytogenetic Testing

• Cytogenetic testing is used to:
  • Study changes in chromosome number and structure in populations.
  • Assess the relationship between chromosomal changes and reproductive characteristics, productivity, vitality, and disease resistance.
  • Identify chromosomal markers for research purposes.
  • Perform pre-implantation and prenatal diagnostics.
  • Determine embryo sex and chromosomal changes.
  • Evaluate mitogen effects on the genome.
  • Generate chromosomal maps.

Variation in Chromosome Sets Between Species

• Different species have varying numbers of chromosomes.

Human Karyotype

• Humans have 46 chromosomes, arranged in 23 pairs.
  • Autosomes: 22 pairs of chromosomes.
  • Sex chromosomes: 1 pair (XX female, XY male).

Chromosomal Abnormalities

• Changes in chromosome number:
  • Polyploidy: A regular increase in the total set of chromosomes (3n, 4n). Often formed as a result of interspecific crossbreeding.
  • Aneuploidy: A change in the number of individual chromosomes:
    • Trisomy (2n+1): An individual has three chromosomes instead of a pair.
    • Monosomy (2n-1): An individual has one chromosome instead of a pair.

Inheritance of Aneuploidy

• Aneuploidy arises from non-disjunction events during meiosis.
  • Meiosis I non-disjunction: Homologous chromosomes fail to separate.
  • Meiosis II non-disjunction: Sister chromatids fail to separate.

Changes in Chromosome Structure

• Deletions: Loss of a chromosome segment.
• Gap: Damage to the protein coat of a chromosome without DNA breakage.
• Translocation: A segment from one chromosome is inserted into another chromosome.
• Inversions: A segment of a chromosome is reversed.

Cytogenetic Methods

• Conventional cytogenetics: Uses staining techniques (Giemza, G-banding, R-banding, C-banding, T-banding, NOR staining).
• Molecular cytogenetics: Uses molecular techniques (FISH, CGH, SKY).

Preparation of Chromosome Preparations

• Direct Method: Chromosome preparations made from tissues with active cell division.
• Indirect Method: Chromosome preparations made from peripheral blood using a mitogen to induce cell division.

Giemza Staining

• Giemza staining: A simple staining technique that reveals the number of chromosomes and major structural changes.

G-banding

• G-banding: A technique that produces distinct dark and light bands on chromosomes.
  • Trypsin treatment: Removes part of the chromosomal proteins.
  • Giemza or acridine orange staining: Produces the banding pattern.
  • A- and T-rich sequences: Stain darkly.
  • C- and G-rich sequences: Stain lightly.

R-banding

• R-banding: A reverse G-banding technique.
  • Heat treatment: Denatures DNA.
  • Giemza or acridine orange staining: Produces a banding pattern where A- and T-rich sequences stain lightly and G- and C-rich sequences stain darkly.

C-banding

• C-banding: A technique that stains constitutive heterochromatin near the centromere.
  • Acid and alkali treatment: Denatures and renatures DNA.
  • Giemza staining: Stains the heterochromatin darkly.
  • Used to identify sex chromosomes and translocations.

T-banding

• T-banding: A modification of R-banding.
  • Heat treatment: Denatures DNA.
  • Stains the telomeric regions of chromosomes: Helps analyze abnormalities in telomeric regions.

NOR (Nucleolar Organizing Region) Staining

• NOR staining: Identifies satellites located on the short arms of acrocentric chromosomes.
  • Silver nitrate staining: Binds to active rRNA genes.

Fluorescence In Situ Hybridization (FISH)

• FISH: A technique that uses fluorescent probes to detect specific chromosome locations.
  • Fluorescent probes: Bind to DNA-target regions based on complementarity.
  • Used to detect changes in chromosome number and structure.

FISH Process Flow

• FISH process:
  • Chromosome preparations or fixed cells are prepared on a slide.
  • DNA is denatured using heat or chemicals.
  • Probes hybridize to the target DNA.
  • Non-hybridized probes are washed away.
  • Results are analyzed using a fluorescence microscope.

This quiz explores the fundamentals of cytogenetics, including the structure and function of chromosomes and chromatin within eukaryotic cells. It covers crucial topics such as nucleosome structure and the differences between euchromatin and heterochromatin. Test your understanding of these essential components of genetics.