PDF(Pubmed)
Abstract：
Targeting, safety, scalability, and storage stability of vectors are still challenges in the field of nucleic acid delivery for gene therapy. Silica-based nanoparticles have been widely studied as gene carriers, exhibiting key features such as biocompatibility, simplistic synthesis, and enabling easy surface modifications for targeting. However, the ability of the formulation to incorporate DNA is limited, which restricts the number of DNA molecules that can be incorporated into the particle, thereby reducing gene expression. Here we use polymerase chain reaction (PCR)-generated linear DNA molecules to augment the coding sequences of gene-carrying nanoparticles, thereby maximizing nucleic acid loading and minimizing the size of these nanocarriers. This approach results in a remarkable 16-fold increase in protein expression six days post-transfection in cells transfected with particles carrying the linear DNA compared with particles bearing circular plasmid DNA. The study also showed that the use of linear DNA entrapped in DNA@SiO2 resulted in a much more efficient level of gene expression compared to standard transfection reagents. The system developed in this study features simplicity, scalability, and increased transfection efficiency and gene expression over existing approaches, enabled by improved embedment capabilities for linear DNA, compared to conventional methods such as lipids or polymers, which generally show greater transfection efficiency with plasmid DNA. Therefore, this novel methodology can find applications not only in gene therapy but also in research settings for high-throughput gene expression screenings.
摘要：
瞄准,安全,可扩展性，和载体的储存稳定性仍然是基因治疗的核酸递送领域的挑战。二氧化硅基纳米粒子作为基因载体已被广泛研究,表现出生物相容性等关键特征，简单化的综合，并能够轻松地进行表面修改以进行瞄准。然而,制剂掺入DNA的能力有限，这限制了可以掺入颗粒中的DNA分子的数量，从而降低基因表达。在这里，我们使用聚合酶链反应（PCR）产生的线性DNA分子来增加携带基因的纳米颗粒的编码序列，从而最大化核酸加载并最小化这些纳米载体的尺寸。与携带环状质粒DNA的颗粒相比，该方法在转染后六天在用携带线性DNA的颗粒转染的细胞中导致蛋白质表达显着增加16倍。该研究还表明,与标准转染试剂相比,使用包埋在DNA@SiO2中的线性DNA导致更有效的基因表达水平。本研究中开发的系统具有简单性，可扩展性，与现有方法相比，提高了转染效率和基因表达，通过改进线性DNA的嵌入能力，与脂质或聚合物等常规方法相比，其通常显示更高的质粒DNA转染效率。因此,这种新方法不仅可以应用于基因治疗，还可以应用于高通量基因表达筛选的研究环境。
