Research output: Contribution to journal › Article › peer-review
A cookbook for DNase Hi-C. / Gridina, Maria; Mozheiko, Evgeniy; Valeev, Emil et al.
In: Epigenetics and Chromatin, Vol. 14, No. 1, 15, 20.03.2021.Research output: Contribution to journal › Article › peer-review
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TY - JOUR
T1 - A cookbook for DNase Hi-C
AU - Gridina, Maria
AU - Mozheiko, Evgeniy
AU - Valeev, Emil
AU - Nazarenko, Ludmila P.
AU - Lopatkina, Maria E.
AU - Markova, Zhanna G.
AU - Yablonskaya, Maria I.
AU - Voinova, Viktoria Yu
AU - Shilova, Nadezhda V.
AU - Lebedev, Igor N.
AU - Fishman, Veniamin
N1 - Publisher Copyright: © 2021, The Author(s). Copyright: Copyright 2021 Elsevier B.V., All rights reserved.
PY - 2021/3/20
Y1 - 2021/3/20
N2 - Background: The Hi-C technique is widely employed to study the 3-dimensional chromatin architecture and to assemble genomes. The conventional in situ Hi-C protocol employs restriction enzymes to digest chromatin, which results in nonuniform genomic coverage. Using sequence-agnostic restriction enzymes, such as DNAse I, could help to overcome this limitation. Results: In this study, we compare different DNAse Hi-C protocols and identify the critical steps that significantly affect the efficiency of the protocol. In particular, we show that the SDS quenching strategy strongly affects subsequent chromatin digestion. The presence of biotinylated oligonucleotide adapters may lead to ligase reaction by-products, which can be avoided by rational design of the adapter sequences. Moreover, the use of nucleotide-exchange enzymes for biotin fill-in enables simultaneous labelling and repair of DNA ends, similar to the conventional Hi-C protocol. These improvements simplify the protocol, making it less expensive and time-consuming. Conclusions: We propose a new robust protocol for the preparation of DNAse Hi-C libraries from cultured human cells and blood samples supplemented with experimental controls and computational tools for the evaluation of library quality.
AB - Background: The Hi-C technique is widely employed to study the 3-dimensional chromatin architecture and to assemble genomes. The conventional in situ Hi-C protocol employs restriction enzymes to digest chromatin, which results in nonuniform genomic coverage. Using sequence-agnostic restriction enzymes, such as DNAse I, could help to overcome this limitation. Results: In this study, we compare different DNAse Hi-C protocols and identify the critical steps that significantly affect the efficiency of the protocol. In particular, we show that the SDS quenching strategy strongly affects subsequent chromatin digestion. The presence of biotinylated oligonucleotide adapters may lead to ligase reaction by-products, which can be avoided by rational design of the adapter sequences. Moreover, the use of nucleotide-exchange enzymes for biotin fill-in enables simultaneous labelling and repair of DNA ends, similar to the conventional Hi-C protocol. These improvements simplify the protocol, making it less expensive and time-consuming. Conclusions: We propose a new robust protocol for the preparation of DNAse Hi-C libraries from cultured human cells and blood samples supplemented with experimental controls and computational tools for the evaluation of library quality.
KW - A549
KW - DNAse I
KW - Genome organization
KW - Hi-C
KW - Human peripheral blood
KW - K562
KW - LNCaP
UR - http://www.scopus.com/inward/record.url?scp=85103169929&partnerID=8YFLogxK
U2 - 10.1186/s13072-021-00389-5
DO - 10.1186/s13072-021-00389-5
M3 - Article
C2 - 33743768
AN - SCOPUS:85103169929
VL - 14
JO - Epigenetics and Chromatin
JF - Epigenetics and Chromatin
SN - 1756-8935
IS - 1
M1 - 15
ER -
ID: 28205303