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Lecture 4
DNA packaging
14
Biology
Professional
08/13/2011

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Term
Histone complexes (name of basic structural unit of chromatin, # of bp/turns per complex, # of bp between complexes
Definition
  • nucleosome
  • 146bp and 1.8 turns around each histone core
  • 20 - 60 bp spacer
Term
DNA packaging heirarchy
Definition
  1. double helix - 2nm 
  2. nucleosome fiber - beads on a string - 10nm
  3. solenoid - 30 nm fiber
  4. solenoid packed in loops and attached to protein scaffold/ nuclear matrix at fixed locations - attachment intervals ~100kb
Term
Specialized histones/ Histone code
Definition
  • H3 + H2A: confer specific characteristics - genomic DNA
  • H3 + H4: can be modefied by chemical changes 
    • Post-translational modifications chng properties of nucleosomes that contain them 
  • histone code: pattern of major and specialized histone types and their modifications
    • Varies from cell type to cell type, specifies packaging, and allows DNA to be accessible to regulatory molecules that determine gene expression
Term
Types of histone modifications
Definition
  • acetylation
  • methylation
  • ubiquitination
  • sumoylation
  • phosphorylation 
Term
Is DNA packaged the same in all cell types?
Definition
  • No! Sperm cells highly specialized
  • Histone proteins replaced with specialized protamines at late spermatogenesis
    • Protamines are argenine rich and enable tighter packaging essential for sperm head condensation and DNA stabilization 
Term
How does chromatin condensation chng with cell cycle?
Definition
  • interphase - decondensed solenoid loops
  • condenses as mitosis begins
  • fully condensed at metaphase
  • decondenses again toward interphase 
Term
Telomeres and Centromeres
Definition
  • Telomeres: seal chromosome ends preventing fusion with other chromosomes (TTAGGG repeats)
  • Centromeres: specialized region of DNA (constriction along chromosome) - During cell division provides site at which spindle apparatus is anchored - essential for correct chromosome segregation 
Term
Chromosome Preparation Procedure
Definition
  1. blood sample (WBC)
  2. cell culture - PHA (phytohemaglutinin) induce mitosis (condensation)
  3. treat with colchicine (dissolves spindle apparatus) and hypotonic soln.  
  4. centrifuge
  5. fix cells (methanol + acetic acid)
  6. spread cells on slides, stain, microsopy
Term
types of chromosome banding methods
Definition
  • G-banding - Giemsa - stains AT rich regions - most common in karyotyping
  • c-banding - distinguish heterochromatin regions - favors areas like centromeres
  • q-banding - quinacrine - fluorescent, similar to G bands
  • r-banding - reverse of C banding, stains non-centromeric regions
  • Reverse DAPI banding - similar to G banding
  • Ag-NOR active nucleolar organising region
Term
names for diff centromere position
Definition
  • metacentric - 1/2
  • submetacentric - 1/3
  • acrocentric - small piece of DNA on one side of centromere 
Term
chromosome nomenclature
Definition
  • p = short arm (petite)
  • q = long arm (p+q = 1)
  • cen = centromere
  • ter = terminal/ telomere 
Term
dark vs light bands in G-banding
Definition
  • Dark bands - more condensed, A-T rich regions, heterochromatin, gene poor regions
  • Light bands - less condensed, G-C rich regions, euchromatin, gene rich regions 
Term
indications for karyotyping
Definition
  • Probs in early growth + development (malformation, short stature, retardation
  • stillbirth + neonatal death
  • fertility probs (amenorrhea, hx of infertility, recurrent miscarriage)
  • family hx of chromosome abnormality in 1st deg. relative
  • neoplasia - cancer
  • pregnancy > 35 yrs of age - fetal chromosome analysis 
Term
FISH
Definition
  1. chromosome prep on slide
  2. denature DNA in situ
  3. label by incorp. nucleotides with attached flourophore or single probe for specific sequence
  4. allow to anneal, expose to UV, and visualize in situ 
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