Editing Nucleosome (section)

==Nucleosome assembly ''in vitro''==
[[Image:Nucleosome structure.png|right|thumb|Diagram of nucleosome assembly]]
Nucleosomes can be assembled ''[[in vitro]]'' by either using purified native or recombinant histones.<ref>{{cite journal | vauthors = Hayes JJ, Lee KM | title = In vitro reconstitution and analysis of mononucleosomes containing defined DNAs and proteins | journal = Methods | volume = 12 | issue = 1 | pages = 2–9 | date = May 1997 | pmid = 9169189 | doi = 10.1006/meth.1997.0441 | doi-access = free }}</ref><ref>{{cite book | vauthors = Dyer PN, Edayathumangalam RS, White CL, Bao Y, Chakravarthy S, Muthurajan UM, Luger K | title = Chromatin and Chromatin Remodeling Enzymes, Part A | chapter = Reconstitution of nucleosome core particles from recombinant histones and DNA | series = Methods in Enzymology | volume = 375 | pages = 23–44 | year = 2004 | pmid = 14870657 | doi = 10.1016/s0076-6879(03)75002-2 | isbn = 9780121827793 }}</ref> One standard technique of loading the DNA around the histones involves the use of salt [[Kidney dialysis|dialysis]]. A reaction consisting of the histone octamers and a naked DNA template can be incubated together at a salt concentration of 2 M. By steadily decreasing the salt concentration, the DNA will equilibrate to a position where it is wrapped around the histone octamers, forming nucleosomes.  In appropriate conditions, this reconstitution process allows for the nucleosome positioning affinity of a given sequence to be mapped experimentally.<ref>{{cite journal | vauthors = Yenidunya A, Davey C, Clark D, Felsenfeld G, Allan J | title = Nucleosome positioning on chicken and human globin gene promoters in vitro. Novel mapping techniques | journal = Journal of Molecular Biology | volume = 237 | issue = 4 | pages = 401–414 | date = April 1994 | pmid = 8151701 | doi = 10.1006/jmbi.1994.1243 }}</ref>

=== Disulfide crosslinked nucleosome core particles ===
A recent advance in the production of nucleosome core particles with enhanced stability involves site-specific [[disulfide]] crosslinks.<ref>{{cite journal | vauthors = Frouws TD, Barth PD, Richmond TJ | title = Site-Specific Disulfide Crosslinked Nucleosomes with Enhanced Stability | journal = Journal of Molecular Biology | volume = 430 | issue = 1 | pages = 45–57 | date = January 2018 | pmid = 29113904 | pmc = 5757783 | doi = 10.1016/j.jmb.2017.10.029 }}</ref> Two different crosslinks can be introduced into the nucleosome core particle. A first one crosslinks the two copies of [[Histone H2A|H2A]] via an introduced cysteine (N38C) resulting in [[histone octamer]] which is stable against H2A/H2B dimer loss during nucleosome reconstitution. A second crosslink can be introduced between the H3 N-terminal histone tail and the nucleosome DNA ends via an incorporated convertible nucleotide.<ref>{{cite book | vauthors = Ferentz AE, Verdine GL |chapter=The Convertible Nucleoside Approach: Structural Engineering of Nucleic Acids by Disulfide Cross-Linking |pages=14–40 |doi=10.1007/978-3-642-78666-2_2 | veditors = Eckstein F, Lilley DM |title=Nucleic Acids and Molecular Biology |volume=8 |year=1994 |isbn=978-3-642-78668-6 }}</ref> The DNA-histone octamer crosslink stabilizes the nucleosome core particle against DNA dissociation at very low particle concentrations and at elevated salt concentrations.