Encapsulation of Nucleic Acids into Giant Unilamellar Vesicles by Freeze-Thaw: a Way Protocells May Form.
Protocells are believed to consist of a lipid membrane and encapsulated nucleic acid. As the lipid membrane is impermeable to macromolecules like nucleic acids, the processes by which nucleic acids become encapsulated inside lipid membrane compartments are still unknown. In this paper, a freeze-thaw method was modified and applied to giant unilamellar vesicles (GUVs) and deoxyribonucleic acid (DNA) in mixed solution resulting in the efficient encapsulation of 6.4 kb plasmid DNA and similar length linear DNA into GUVs. The mechanism of encapsulation was followed by observing the effect of freeze-thaw temperatures on GUV morphological change, DNA encapsulation and ice crystal formation, and analyzing their correlation. Following ice crystal formation, the shape of spherical GUVs was altered and membrane integrity was damaged and this was found to be a necessary condition for encapsulation. Heating alone had no effects on DNA encapsulation, but was helpful for restoring the spherical shape and membrane integrity of GUVs damaged during freezing. These results suggested that freeze-thaw could promote the encapsulation of DNA into GUVs by a mechanism: the vesicle membrane was breached by ice crystal formation during freezing, DNA entered into damaged GUVs through these membrane gaps and was encapsulated after the membrane was resealed during the thawing process. The process described herein therefore describes a simple way for the encapsulation of nucleic acids and potentially other macromolecules into lipid vesicles, a process by which early protocells might have formed.
State Key Laboratory of Ultrasound Engineering in Medicine Co-Founed by Chongqing and the Ministry of Science and Technology, Chongqing Key Laboratory of Biomedical Engineering, College of Biomedical Engineering, Chongqing Medical University, P. O. Box 153, No.1 Yixueyuan Road, Yuzhong District, Chongqing, 400016, People's Republic of China.
Hai
http://orcid.org/0000-0001-9818-7149
H
Qiao
State Key Laboratory of Ultrasound Engineering in Medicine Co-Founed by Chongqing and the Ministry of Science and Technology, Chongqing Key Laboratory of Biomedical Engineering, College of Biomedical Engineering, Chongqing Medical University, P. O. Box 153, No.1 Yixueyuan Road, Yuzhong District, Chongqing, 400016, People's Republic of China.
Na
N
Hu
State Key Laboratory of Ultrasound Engineering in Medicine Co-Founed by Chongqing and the Ministry of Science and Technology, Chongqing Key Laboratory of Biomedical Engineering, College of Biomedical Engineering, Chongqing Medical University, P. O. Box 153, No.1 Yixueyuan Road, Yuzhong District, Chongqing, 400016, People's Republic of China.
Jin
J
Bai
State Key Laboratory of Ultrasound Engineering in Medicine Co-Founed by Chongqing and the Ministry of Science and Technology, Chongqing Key Laboratory of Biomedical Engineering, College of Biomedical Engineering, Chongqing Medical University, P. O. Box 153, No.1 Yixueyuan Road, Yuzhong District, Chongqing, 400016, People's Republic of China.
Lili
L
Ren
State Key Laboratory of Ultrasound Engineering in Medicine Co-Founed by Chongqing and the Ministry of Science and Technology, Chongqing Key Laboratory of Biomedical Engineering, College of Biomedical Engineering, Chongqing Medical University, P. O. Box 153, No.1 Yixueyuan Road, Yuzhong District, Chongqing, 400016, People's Republic of China.
Qing
Q
Liu
State Key Laboratory of Ultrasound Engineering in Medicine Co-Founed by Chongqing and the Ministry of Science and Technology, Chongqing Key Laboratory of Biomedical Engineering, College of Biomedical Engineering, Chongqing Medical University, P. O. Box 153, No.1 Yixueyuan Road, Yuzhong District, Chongqing, 400016, People's Republic of China. lqfang06@163.com.
Liaoqiong
L
Fang
State Key Laboratory of Ultrasound Engineering in Medicine Co-Founed by Chongqing and the Ministry of Science and Technology, Chongqing Key Laboratory of Biomedical Engineering, College of Biomedical Engineering, Chongqing Medical University, P. O. Box 153, No.1 Yixueyuan Road, Yuzhong District, Chongqing, 400016, People's Republic of China. wangzb@cqmu.edu.cn.
Zhibiao
Z
Wang
D058045:Artificial Cells / Q000166:cytology / Q000378:metabolism*; D005615:Freezing; D009696:Nucleic Acids / Q000378:metabolism*; D000069497:Origin of Life; D053835:Unilamellar Liposomes / Q000378:metabolism*
D009696:Nucleic Acids; D053835:Unilamellar Liposomes
Deoxyribonucleic acid; Encapsulation; Freeze-thaw; Giant unilamellar vesicles; Protocells