Development of novel gene delivery system

阳离子脂质囊泡介导的基因转移已成为体外基因传递的常用方法。然而，转染效率经常受到血清的影响。我们之前报道的DDAB（二甲基二十八烷基溴化铵）脂质囊泡介导的基因转移也存在同样的问题。为了克服这一障碍，我们开发了一种新的转染载体，使用蛋白修饰的DDAB脂质囊泡。在之前的研究中，游离鱼精蛋白被简单地添加到DNA/脂质复合物中，而在本方法中，鱼精蛋白被化学偶联到硬脂酸中，并被掺入到DDAB脂质囊泡中。用该方法转染培养细胞，在含10%血清的培养基中，基因转移无明显抑制。蛋白蛋白修饰的DDAB脂质囊泡也可以利用复制缺陷逆转录病毒载体在血清存在下提高病毒转导效率。此外，这些囊泡可以在体内有效地转移禽类胚胎的基因。这些结果表明，该方法可用于转基因动物的生产和基因治疗。

Cationic lipid vesicle-mediated gene transfer has become common for in vitro gene delivery. However, the transfection efficiency is often impaired by serum. DDAB (dimethyldioctadecyl ammonium bromide) lipid vesicle-mediated gene transfer, which we previously reported, has the same problem. To overcome this obstacle, we develop a novel transfection vehicle using protamine-modified DDAB lipid vesicles. While free protamine was simply added to the DNA/lipid complex in the previous study, in the present method the protamine is chemically conjugated to stearic acid and incorporated into DDAB lipid vesicles. Gene transfer was not significantly inhibited in 10% serumcontaining medium by this method for the transfection of cultured cells. Protaminemodified DDAB lipid vesicles also enhanced virus transduction efficiency in the presence of serum using a replication'defective retroviral vector. Furthermore, the vesicles allowed efficient gene transfer for avian embryos in vivo. These results indicate that the method is useful for the production oftransgenic animals and gene therapy.