Freezing shortens the lifetime of DNA molecules under tension.
Freezing shortens the lifetime of DNA molecules under tension.
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DOI：
10.1007/s10867-017-9466-3
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发表时间：
2017-12
影响因子：
1.8
通讯作者：
Hsu IC
中科院分区：
文献类型：
--
作者：
Chung WJ;Cui Y;Chen CS;Wei WH;Chang RS;Shu WY;Hsu IC
DNA样本通常被冷冻保存。然而，冷冻会损害DNA分子的完整性。考虑到DNA分子在纳米技术中的广泛应用，在分子水平上改变DNA完整性可能会导致不良后果。然而，冷冻对DNA完整性的影响尚未得到充分探讨。为了研究冷冻对DNA完整性的影响，使用光学镊子研究了冷冻和非冷冻噬菌体lambda DNA样本。在DNA分子上施加张力（5-35 pN）来模拟DNA与其他生物分子之间的机械相互作用。DNA分子的完整性是通过测量单个DNA分子在张力下断裂的时间来评估的。平均寿命由最大似然估计确定，方差通过自举模拟获得。在5 pN的力作用下，冷冻样品的平均寿命为44.3 min， 95%可信区间（CI）在36.7 ~ 53.6 min之间，而非冷冻样品的平均寿命为133.2 min （95% CI: 97.8 ~ 190.1 min）。在15 pN压力下，平均生存时间分别为10.8 min （95% CI: 7.6-12.6 min）和78.5 min （95% CI: 58.1-108.9 min）。冷冻DNA分子的寿命大大缩短，这意味着冷冻损害了DNA的完整性。此外，我们发现减少的DNA结构完整性不能恢复使用常规的结扎过程。这些结果表明，冷冻可以改变DNA分子的结构完整性。
DNA samples are commonly frozen for storage. However, freezing can compromise the integrity of DNA molecules. Considering the wide applications of DNA molecules in nanotechnology, changes to DNA integrity at the molecular level may cause undesirable outcomes. However, the effects of freezing on DNA integrity have not been fully explored. To investigate the impact of freezing on DNA integrity, samples of frozen and non-frozen bacteriophage lambda DNA were studied using optical tweezers. Tension (5–35 pN) was applied to DNA molecules to mimic mechanical interactions between DNA and other biomolecules. The integrity of the DNA molecules was evaluated by measuring the time taken for single DNA molecules to break under tension. Mean lifetimes were determined by maximum likelihood estimates and variances were obtained through bootstrapping simulations. Under 5 pN of force, the mean lifetime of frozen samples is 44.3 min with 95% confidence interval (CI) between 36.7 min and 53.6 min while the mean lifetime of non-frozen samples is 133.2 min (95% CI: 97.8–190.1 min). Under 15 pN of force, the mean lifetimes are 10.8 min (95% CI: 7.6–12.6 min) and 78.5 min (95% CI: 58.1–108.9 min). The lifetimes of frozen DNA molecules are significantly reduced, implying that freezing compromises DNA integrity. Moreover, we found that the reduced DNA structural integrity cannot be restored using regular ligation process. These results indicate that freezing can alter the structural integrity of the DNA molecules.
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