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2016 ; 114
(ä): 21.33.1-21.33.11
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Molecular Dissection of Chromatin Maturation via Click Chemistry
#MMPMID27038388
Yildirim O
; Kingston RE
Curr Protoc Mol Biol
2016[Apr]; 114
(ä): 21.33.1-21.33.11
PMID27038388
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DNA synthesis and chromatin assembly are the two most critical processes of
eukaryotic cell division. It is well known that their coordination is tightly
regulated. Although the interplay between DNA and its higher-order chromatin
state is integral for many processes, including cell survival and genome
stability, little is known about the re-establishment of chromatin structure
during the cell cycle. Moreover, the extent to which the fidelity of the newly
synthesized chromatin plays a role in the maintenance of cellular identity is
still under debate. Here, we present a novel approach to purify nascent chromatin
from the replication fork. In this protocol, we take advantage of click
chemistry, a method that allows efficient conjugation of azide-containing biotin
molecules to ethynyl-labeled nucleic acids. Using this approach, we selectively
enrich biotin-nucleic acid conjugates via streptavidin affinity purification to
pull down and assess chromatin states as well as chromatin-bound complexes from
newly replicated DNA fragments.
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