Transient transfection of primary cultured hepatocytes using CaPO4/DNA precipitation.
Transient transfection of primary cultured hepatocytes using CaPO4/DNA precipitation.
复制标题
使用 CaPO4/DNA 沉淀瞬时转染原代培养的肝细胞。
作者：
F. Gaunitz;M. Papke;R. Gebhardt
我们提出了一种易于复制的非增殖性原代培养肝细胞瞬时转染法。使用经典的CaPO4/DNA沉淀法的改进，该方法是一种廉价的替代其他方法的方法，这些方法通常繁琐、昂贵、难以复制或对原代肝细胞有害。因为一次只需要0.5×10(6)个细胞，所以一次肝制剂中可以引入几个报告基因。在我们的方案中，不同的质粒可以被导入一个细胞。通过这种方法，可以检测顺式和反式的相互作用，并且可以标准化报告基因的表达以提高转染率。此外，我们描述了使用荧光素酶基因和lacZ基因用两种不同的报告基因载体进行的转染实验的细节。这些结果可能会对其他关注改进转基因实验时机的小组有所帮助。
We present a detailed protocol for the transient transfection of non-proliferating primary cultured hepatocytes that is easily reproducible. Using a modification of the classical CaPO4/DNA precipitation method, this protocol is an inexpensive alternative to other methods that are often cumbersome, expensive, difficult to reproduce or harmful to primary hepatocytes. Because only 0.5 x 10(6) cells are needed for a single transfection experiment, several reporter genes can be introduced into hepatocytes of a single liver preparation. With our protocol, different plasmids can be introduced into one cell. In this way, cis-trans interactions can be examined and reporter gene expression can be normalized for transfection efficiency. Furthermore, we describe details of a transfection experiment with two different reporter gene vectors using a luciferase gene and a lacZ gene. The results presented may be helpful to other groups concerned with improved timing of transfection experiments.
影响因子：
2.7
作者：
S. Nordeen
通讯作者：
S. Nordeen
通过在培养皿中与 β-半乳糖苷酶底物一起孵育细胞来进行 DNA 转染的简单测定。
影响因子：
2.7
作者：
Lim,K;Chae,CB
通讯作者：
Chae,CB