平板离心提高了聚乙烯亚胺转染和慢病毒感染的效率。

IF 2.2 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods Pub Date : 2024-09-30 DOI:10.1016/j.jviromet.2024.115039

Shaozhe Yang , Qingwei Zhang , Yuan Zhuang , Junfeng Li , Xiuhong Fu

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引用次数: 0

摘要

目的：提出一种基于平板离心的高效、可重复和稳定的转基因技术，该技术尤其适用于聚乙烯亚胺（PEI）转染和慢病毒感染：我们优化了可能影响转染效率的多个因素，如 PEI 或 DNA 的用量、稀释 PEI 或 DNA 的工作液缓冲液、PEI/DNA 复合物的孵育时间以及转染时间：结果：平板离心使基于 PEI 的转染效率提高了 5.46 倍，同时保持了细胞存活率。此外，平板离心使每个基因组中病毒基因的平均拷贝数增加了 4.96 倍。平板离心改变了 PEI/DNA 复合物或慢病毒的空间排列，增加了这些复合物或病毒与靶细胞接触的机会，最终提高了转染或感染效率：我们提出了一种基于平板离心的转染或慢病毒感染方案，适用于原代细胞或干细胞的基因改造。

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Plate centrifugation enhances the efficiency of polyethylenimine-based transfection and lentiviral infection

Purpose

To propose an efficient, reproducible, and consistent transgenic technology based on plate centrifugation, which is particularly useful for polyethylenimine (PEI) transfection and lentiviral infection.

Methods

We optimized multiple factors that could contribute to transfection efficiency, such as the dosage of the PEI or DNA, the working solution buffer used for diluting the PEI or DNA, the incubation time for the PEI/DNA complexes, and the transfection time.

Results

Plate centrifugation led to a 5.46-fold increase in the transfection efficiency of PEI-based transfection while maintaining the cell survival rate. Moreover, the average copy number of viral genes in each genome increased 4.96-fold with plate centrifugation. Plate centrifugation alters the spatial arrangement of the PEI/DNA complexes or lentiviruses, increasing the chances of these complexes or viruses coming into contact with target cells, ultimately resulting in improved transfection or infection efficiency.

Conclusions

We present a protocol based on plate centrifugation for transfection or lentiviral infection that is suitable for genetic modification of primary cells or stem cells.

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来源期刊

Journal of virological methods 医学-病毒学

CiteScore

5.80

自引率

0.00%

发文量

209

审稿时长

41 days

期刊介绍： The Journal of Virological Methods focuses on original, high quality research papers that describe novel and comprehensively tested methods which enhance human, animal, plant, bacterial or environmental virology and prions research and discovery. The methods may include, but not limited to, the study of: Viral components and morphology- Virus isolation, propagation and development of viral vectors- Viral pathogenesis, oncogenesis, vaccines and antivirals- Virus replication, host-pathogen interactions and responses- Virus transmission, prevention, control and treatment- Viral metagenomics and virome- Virus ecology, adaption and evolution- Applied virology such as nanotechnology- Viral diagnosis with novelty and comprehensive evaluation. We seek articles, systematic reviews, meta-analyses and laboratory protocols that include comprehensive technical details with statistical confirmations that provide validations against current best practice, international standards or quality assurance programs and which advance knowledge in virology leading to improved medical, veterinary or agricultural practices and management.