Chromatin Lecture 1 2013.pdf

Kinetics of digesting nucleohistone with micrococcal

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-- kinetics of digesting nucleohistone with micrococcal nuclease (Mnase) -- rapidly digested fraction and a slow or resistant fraction -- concluded that ~50% of the DNA was protected by histones 1973-- Hewish and Burgoyne BBRC 52, 504-510 -- rat liver nuclein incubated with Ca +2 / Mg +2 -- endogenous nucleases produced breaks in DNA, ladder of DNA fragments. -- MWs were multiples of the smallest species nuclease sensitive nuclease resistant
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Early Structure of Chromatin Fibre 1974 Noll Nucleic Acids Research 1, 1573-1578 -- “in situ” digestion of chromatin by adding exogenous Ca +2 /Mg +2 - dependent nuclease -- estimated DNA content of 205 bp -- from sedimentation density, protein mass = 1.3 X DNA mass
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Early Structure of Chromatin Fibre 1974-- Kornberg and Thomas Science 184, 865-868 -- extracted histones from chromatin with high salt -- observed a histone subunit without DNA -gel filtration, denaturing PAGE and analytical centrifugation data -- concluded: H3/H4 tetramer of 54 kDa, H2A/H2B oligomer (later shown to be a dimer) -- reconstituted H3/H4 tetramer with DNA, low angle x-ray scattering, looked like chromatin
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Early Structure of Chromatin Fibre 1973 Olins and Olins ASCB meeting -- swollen, ruptured rate liver nuclei, chicken erythrocyte, calf thymus -- fixed, spun down onto carbon-coated EM grids, negative stained -- observed 70Å diameter ν - bodies Olins and Olins 1974 Science 183, 330-332 Chris Woodcock submitted similar EM results at the same time to Science. Here is a quote from the reviewer of the manuscript (the paper was rejected, obviously): “... A eukaryotic chromosome made out of self— assembling 70 Å units, which could perhaps be made to crystallize, would necessitate require rewriting our textbooks on cytology and genetics! I have never read such a naive paper supporting to be of such fundamental significance. Definitely it should not be published anywhere."
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Fast forward 20 years High resolution model of the nucleosome
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~6 nm 11 nm What is the compaction ratio of a nucleosome core particle? How much does a nucleosome change the “effective” length of DNA?
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~6 nm 11 nm What is the compaction ratio (CR) of a nucleosome core particle? CR = Length of DNA in the core particle / dimension of core particle How much does a nucleosome change the “effective” length of DNA?
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~6 nm 11 nm What is the compaction ratio (CR) of a nucleosome core particle?
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