Differences in DNA

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Differences in DNA

• Heterochromatin vs. Euchromatin
• Heterochromatin is DNA which tends to be highly
  compacted and dark staining.
• Euchromatin is not so compacted or dark.
• The number of genes in heterochromatin is
  generally small relative to euchromatin.
• Heterochromatin lacks genes or they are inactive
• Much heterochromatin is found in certain
  structural parts of the chromosomes centromeres
  and telomeres. Also, much of Y chromosome.
• Move euchromatin to an area next to
  heterochromatin and it becomes heterochromatin
  position effect.

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Chromosome structure
Arm
http//www.med.uiuc.edu/m1/genetics/images/webun1/
Chromosome.gif medic.med.uth.tmc.edu/.../
cellbio/hist-01.htm
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More on Differences in DNA

• Base sequences are obviously different from one
  organism to another, but overall DNA composition
  can differ as well.
• In most eukaryotic organisms, DNA composition is
  not uniform across all the DNA in the cell
  patches within the same cell where DNA
  composition is distinct from other regions.

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Composition of DNA GC
There is always equal s of A and T, and G and C,
but the percentage of GC pairs and AT pairs
can be different among different organisms.
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Basepairs held together by H-bonds

• T-A base pairs are held together by 2 H-bonds
• G-C base pairs are held together by 3 H-bonds.
• Therefore G-C pairs require slightly more energy
  to separate.

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Measuring GC hyperchromic shift
As DNA melts, becomes SS, absorbs more UV at
260 nm. Because G-C pairs have 3 H-bonds instead
of two, DNA with more GC is more stable, melts
at higher temperature (blue).
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Satellite DNA

• In prokaryotes, the GC base pairs is pretty
  much averaged out over the entire DNA not so
  with eukaryotes.
• Density gradient ultracentrifugation can also be
  used to determine GC.
• GC pairs are denser than AT, migrate to a lower
  location (greater density) in the gradient.
• Fragmented eukaryotic DNA showed something odd

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Satellite DNA
When the DNA was analyzed, a portion has a lower
GC than the rest of the DNA, producing a
satellite band. How could a portion of DNA
have a different composition than the rest?
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Repeated sequences

• If a section of DNA with a GC composition
  different from the rest of the DNA is repeated
  many times, DNA fragments from these regions of
  DNA would behave differently during the
  centrifugation.

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Study of the Composition of DNA using DNA
renaturation kinetics

• Break DNA into random fragments.
• Denature with heat (melt).
• Cool, allow strands to find their complements and
  go from ss to ds again (anneal/renature).
• Follow entire process using UV light absorption
  at 260 nm
• as DNA goes from ss to ds, Abs decreases.

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Renaturation kinetics

• Kinetics study of the rate of change.
• Major Point 1 the more copies of the
  complementary strands there are, the less time
  they will take to
• find each other
• the more DNA,
• the faster the process.

In this fig., 2 different amounts of DNA from the
SAME organism.
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Renaturation kinetics-2

• Major Point 2
• Given equal amounts
• (same mass) of DNA,
• the bigger the total genome
• of the organism, the slower
• the renaturation.
• If the genome is bigger, and the amounts of DNA
  used in the experiment are the same, the organism
  with the bigger genome will have fewer copies of
  the complementary fragments, so annealing will
  take longer (see point 1).

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Understanding genome size
Imagine you have 20 playing cards. In one
instance, you have these 5 cards, another 5 cards
exactly the same, and 2 more sets of the Ace thru
10 but of diamonds. ltDeck 1gt In the second
instance, you have ace thru 5 of hearts and also
of diamonds. ltDeck 2gt
In which case will you match up pairs of hearts
and diamonds most quickly? The Deck 1 gets
matched up quicker.
http//www.skydiveelsinore.com/calendar/images/pla
ying-cards-spread.jpg
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Cot curves Studying renaturation of DNA
The amount of DNA affects the rate at which DNA
fragments renature. To avoid the problem of
comparing samples with different amounts of DNA,
the change in ss DNA is graphed vs.the initial
DNA concentration (Co) x the time (t)
Cot Y-axis is the fraction or percent of the DNA
that is ss (experiment starts by denaturing the
DNA). X-axis is Cot which is a Log scale.
www.cas.muohio.edu/.../gene2000/ lect7/fig9p8c.jpg
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Satellite DNA and Cot curves
When human DNA was analyzed this way, this was
the result
Remember the card deck experiment when there is
only one of each card in the deck, they take
longer to match up. So DNA that anneals quickly
must be in multiple copies
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Cot curves and satellite DNA
Categories variable among different organisms.
Highly repetitive DNA, many complements, find
each other quickly. Single copy (unique sequence)
much slower.
http//www.ndsu.nodak.edu/instruct/mcclean/plsc431
/eukarychrom/cot2.gif
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Types of DNA

• Highly repetitive DNA 5-45 of DNA depending on
  species. In humans
• ALU family contains Alu I site. 300 bp long,
  appears 500,000 times, dispersed. 5 of DNA.
• SINEs short interspersed elements
• transposable
• Alpha satellite DNA tandem repeats of 170 bp
  occur 5,000-15,000 times make up part of
  centromere. 6
• L1 family (in humans), example of LINEs
• Long interspersed elements
• transposable

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DNA in fewer copies

• Moderately (middle) repetitive DNA
• Tandem or interspersed repeats
• VNTRs, good for DNA fingerprinting
• Variable number tandem repeats
• 15 100 bp long, between or within genes
• Dinucleotide repeats (CA)N, also good for
  forensic work
• in maize and yeasts transposons in large
  numbers.
• genes for rRNA, ribosomal proteins, histones

• Unique, single copy typically 30-75 of DNA in
  most eukaryotes.

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All your DNA codes for proteins? Sorry, not
close

• Only 4 codes for proteins, in 30,000 genes
• 96 of DNA includes
• Introns, junk DNA within and around genes.
• Genes coding for rRNA and tRNA
• Junk DNA called repetitive sequences
• Pseudogenes have sequences that look like genes
  but are never expressed, dont work.
• We are related to everything else
• Our genes look like those from chimpanzees,
  bacteria.