Double-strand breaks in genome-sized DNA caused by mechanical stress under mixing: Quantitative evaluation through single-molecule observation

[Display omitted] •Double-strand breaks (DSBs) in giant DNA molecules caused by mixing were quantitatively evaluated.•The probability of DSBs is quite high at the very beginning of mixing.•A novel technique for mixing to significantly decrease DSBs is proposed. It is becoming increasingly apparent t...

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Bibliographic Details
Published in:Chemical physics letters 2018-06, Vol.701, p.81-85
Main Authors: Kikuchi, Hayato, Nose, Keiji, Yoshikawa, Yuko, Yoshikawa, Kenichi
Format: Article
Language:English
Subjects:
Double-strand breaks
Genome-sized DNA
Mechanical agitation
Mixing procedure
Single-molecule observation
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Summary:[Display omitted] •Double-strand breaks (DSBs) in giant DNA molecules caused by mixing were quantitatively evaluated.•The probability of DSBs is quite high at the very beginning of mixing.•A novel technique for mixing to significantly decrease DSBs is proposed. It is becoming increasingly apparent that changes in the higher-order structure of genome-sized DNA molecules of more than several tens kbp play important roles in the self-control of genome activity in living cells. Unfortunately, it has been rather difficult to prepare genome-sized DNA molecules without damage or fragmentation. Here, we evaluated the degree of double-strand breaks (DSBs) caused by mechanical mixing by single-molecule observation with fluorescence microscopy. The results show that DNA breaks are most significant for the first second after the initiation of mechanical agitation. Based on such observation, we propose a novel mixing procedure to significantly decrease DSBs.
ISSN:0009-2614
1873-4448
DOI:10.1016/j.cplett.2018.04.042