基于核定位序列-DNA四面体复合纳米结构的细胞核成像研究

赵彦; 李凡; 郭琳洁; 代江兵; 邢淑; 王丽华

doi:10.11889/j.0253-3219.2017.hjs.40.050501

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基于核定位序列-DNA四面体复合纳米结构的细胞核成像研究

核物理、交叉学科研究 | 更新时间：2021-05-19

- 基于核定位序列-DNA四面体复合纳米结构的细胞核成像研究
  封面论文
- The nuclei imaging of nucleus localization signal-DNA tetrahedron composite nanostructures
- 核技术 2017年40卷第5期文章编号：050501
- 作者机构：
  
  1.中国科学院上海应用物理研究所物理生物学研究室中国科学院界面物理与技术重点实验室嘉定园区上海 201800
  2.中国科学院大学北京 100049
- 作者简介：
  
  [ "赵彦, 女, 1990年出生, 2012年毕业于山东师范大学, 现为博士研究生, 研究领域为DNA纳米结构的生物学应用", "ZHAO Yan, female, born in1990, graduated from Shandong Normal University in 2012, doctoral student, focusing on the biological applications of DNA nanostructures" ]
  王丽华, E-mail:wanglihua@sinap.ac.cn WANG Lihua, E-mail: wanglihua@sinap.ac.cn
- 基金信息：
  
  国家自然科学基金(No.11575278);国家自然科学基金(No.21675167)
- DOI：10.11889/j.0253-3219.2017.hjs.40.050501
  中图分类号： TL99
- 收稿日期：2017-02-14，
  
  修回日期：2017-02-27，
  
  纸质出版日期：2017-05-10
- 稿件说明：
移动端阅览
赵彦, 李凡, 郭琳洁, 等. 基于核定位序列-DNA四面体复合纳米结构的细胞核成像研究[J]. 核技术, 2017,40(5):050501

Yan ZHAO, Fan LI, Linjie GUO, et al. The nuclei imaging of nucleus localization signal-DNA tetrahedron composite nanostructures[J]. Nuclear techniques, 2017, 40(5): 050501
赵彦, 李凡, 郭琳洁, 等. 基于核定位序列-DNA四面体复合纳米结构的细胞核成像研究[J]. 核技术, 2017,40(5):050501 DOI： 10.11889/j.0253-3219.2017.hjs.40.050501.

Yan ZHAO, Fan LI, Linjie GUO, et al. The nuclei imaging of nucleus localization signal-DNA tetrahedron composite nanostructures[J]. Nuclear techniques, 2017, 40(5): 050501 DOI： 10.11889/j.0253-3219.2017.hjs.40.050501.

摘要

基因治疗的关键是如何高效转运基因进入细胞核并发挥作用，实时观察脱氧核糖核酸（Deoxyribonucleicacid，DNA）在细胞核中的分布对于了解基因治疗的效果非常重要。其中，载体的安全性、运输效率等是基因治疗的关键影响因素。本工作构建了核定位序列（Nuclear Localization Signal，NLS）与DNA四面体的复合结构，提高了细胞核摄取DNA四面体的效率。利用“点击”化学反应将NLS与DNA四面体进行共价连接，解决了简单混合带来的稳定性差等问题。进一步研究了不同长度和等电点的NLS序列在与DNA四面体连接时的效率，发现与经典的NLS

相比，等电点为4.84的NLS

与DNA的非特异性结合显著降低，连接反应的效率从37.3%提高到72.3%。经高效液相色谱（High Performance Liquid Chromatography，HPLC）分离纯化后的NLS

-DNA和NLS

-DNA均可以靶向细胞核，说明改变NLS的长度和等电点后仍保持了很好的活性。该研究改善了正电NLS与负电DNA之间由于非特异性结合容易形成沉淀的问题，为临床基因治疗奠定了基础。

Abstract

Background

Efficient translocation of nucleic acid drugs into nucleus is a key factor in gene therapy. Therefore

real-time observation of the distribution of deoxyribonucleic acid (DNA) in the nucleus is very important to understand the effect of gene therapy. Beforehand

choosing a safe and efficient carrier for translocating DNA into nucleus is critical.

Purpose

This work establishes composite structure of nucleus localization signal (NLS) and DNA tetrahedron

which improving the uptake efficiency of DNA tetrahedron into nucleus.

Methods

Covalently connection NLS and DNA tetrahedron with "click" chemistry solved the problems of poor stability after simple mixing. And we further studied the connecting efficiency of different length and isoelectric point NLS with ssDNA.

Results

Compared with classical NLS

NLS

(of which isoelectric point is 4.84) had a low nonspecific binding. The binding efficiency of NLS

was improved to 72.3% from 37.3%. After purification

both NLS

-DNA and NLS

-DNA could target to cell nucleus

thus verifying that they all possessed good activity.

Conclusion

This research improved nonspecific binding between positively charged NLS and negatively charged DNA

which laid a foundation for clinical gene therapy.

关键词

Keywords

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