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Discovering DNA Packaging into the Nucleosome
Discovering DNA Packaging
Dna Packaging is the folding of DNA to fiit a cell’s limited space. This process differs greatly from Prokaryotic cells to Eukaryotic cells and is much better understood in Eukaryotes. The packaging unit of DNA is called the nucleosome which is basically 8 proteins with 2 DNA strands wrapped around it 2 times. These histone proteins include: H1, H2A, H2B, H3, and H4. When Chromatin (DNA that hasn’t condensed into chromosomes) is put into a low salt solution, it takes the form of “Beads on a string”. It literally looks like those Mardi gras necklaces. When Chromatin is put into a solution with 0.15M KCL, takes the form of the “30-nm fiber” where the DNA now looks like a long thick snake.
DNA packaging has a big problem:
1. The longest human chromosome has 3 X 10^8 base pairs (adenine, cytosine, guanine, thymine). That is equivalent to 10cm. The problem is fitting that much DNA into a cell. A typical cell is 10 micrometers and therefore DNA needs to be compacted 10,000X.
How was the nucleosome (packaging unit of DNA) discovered?
1. The Nuclei was isolated
2. DNAse I (an enzyme which cuts DNA at random locations by breaking phosphodiester bonds) was used to partially digest the DNA (Chop the DNA into fragments).
3. Run the DNA fragments through an agarose gel (which separates the DNA by size)
Result: Scientists found that the DNA fragments were 150- 200 base pairs long (This is weird because DNAse I cuts the DNA at RANDOM places and DNA is VERY long. Because each fragment were about the same length as the next one it means something is blocking the DNA from being randomly chopped up.
Conclusion: This means something is protecting the DNA and that is the histones: H1, H2A, H2B, H3, and H4
To be more precise, the scientists did this:
4. Put in more DNAse so the DNA could be cut up further
5. Run the DNA through Acrylamide gel instead of agarose which provides a better separation
Result: They found that the DNA Fragments are now multiples of 10 – 11 base pairs long. VERY precise.
Conclusion: DNA must be wrapped around the outside of the nucleosome. This also shows that the DNA is bDNA because it is always around 10 base pairs.