Transient Expression of Proteins by Hydrodynamic Gene Delivery in Mice
Transient Expression of Proteins by Hydrodynamic Gene Delivery in Mice
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DOI：
10.3791/51481
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发表时间：
2014-05-01
影响因子：
1.2
通讯作者：
Raper, Jayne
中科院分区：
文献类型：
--
作者：
Kovacsics, Daniella;Raper, Jayne
在体内高效表达转基因基因对研究基因功能和开发疾病治疗方法具有重要意义。近年来，水动力基因传递（HGD）作为一种简单、快速、安全、有效的基因传递方法被广泛应用于啮齿类动物体内。这项技术依靠快速注射大量生理溶液产生的力来增加灌注器官细胞膜的渗透性，从而将DNA传递到细胞中。HGD的主要优点之一是能够使用裸质粒DNA（裸质粒DNA）将转基因引入哺乳动物细胞。使用质粒导入外源基因是最省力、高效率的，而且与病毒载体相反，非常安全。HGD最初用于将基因传递到小鼠体内，现在用于将各种物质，包括寡核苷酸、人工染色体、RNA、蛋白质和小分子传递到小鼠、大鼠体内，并在一定程度上用于其他动物体内。该方案描述了小鼠的HGD，并着重于该方法的三个关键方面，这些方面对于成功执行该过程至关重要：针头正确插入静脉，注射量和递送速度。举例说明了该方法在两个基因的瞬时表达中的应用，这两个基因编码分泌的灵长类特异性蛋白，载脂蛋白L-I （apoli）和触珠蛋白相关蛋白（HPR）。
Efficient expression of transgenes in vivo is of critical importance in studying gene function and developing treatments for diseases. Over the past years, hydrodynamic gene delivery (HGD) has emerged as a simple, fast, safe and effective method for delivering transgenes into rodents. This technique relies on the force generated by the rapid injection of a large volume of physiological solution to increase the permeability of cell membranes of perfused organs and thus deliver DNA into cells. One of the main advantages of HGD is the ability to introduce transgenes into mammalian cells using naked plasmid DNA (pDNA). Introducing an exogenous gene using a plasmid is minimally laborious, highly efficient and, contrary to viral carriers, remarkably safe. HGD was initially used to deliver genes into mice, it is now used to deliver a wide range of substances, including oligonucleotides, artificial chromosomes, RNA, proteins and small molecules into mice, rats and, to a limited degree, other animals. This protocol describes HGD in mice and focuses on three key aspects of the method that are critical to performing the procedure successfully: correct insertion of the needle into the vein, the volume of injection and the speed of delivery. Examples are given to show the application of this method to the transient expression of two genes that encode secreted, primate-specific proteins, apolipoprotein L-I (APOL-I) and haptoglobin-related protein (HPR).