MECHANISMS OF DNA UPTAKE BY MAMMALIAN-CELLS - FATE OF EXOGENOUSLY ADDED DNA MONITORED BY THE USE OF FLUORESCENT DYES
MECHANISMS OF DNA UPTAKE BY MAMMALIAN-CELLS - FATE OF EXOGENOUSLY ADDED DNA MONITORED BY THE USE OF FLUORESCENT DYES
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DOI：
10.1073/pnas.79.2.422
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发表时间：
1982-01-01
期刊：
影响因子：
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通讯作者：
RUDDLE, FH
中科院分区：
文献类型：
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作者：
LOYTER, A;SCANGOS, GA;RUDDLE, FH
DNA 与磷酸钙共沉淀是哺乳动物细胞中常用的基因转移方法。 DNA 与 Ca Pi 形成紧密复合物，并且该复合物中的 DNA 对血清中存在的或外部添加的核酸酶具有抗性。在最佳条件下，几乎所有受体小鼠 Ltk-Aprt- 细胞都会吸收 Ca Pi-DNA 复合物，这是通过 DNA（4'',6-二氨基-2-苯基吲哚二盐酸盐）或钙盐（金霉素）特异的荧光染料确定的。只有一小部分细胞的细胞核中有可检测到的 Ca Pi-DNA 复合物。 Ca Pi-DNA 复合物的吸收高度依赖于形成 Ca Pi-DNA 复合物的 pH 值以及复合物中 DNA 的浓度。
Coprecipitation of DNA with calcium phosphate is a commonly used method of gene transfer in mammalian cells. DNA forms a tight complex with Ca Pi and DNA in this complex is resistant to nucleases present in serum or added externally. Under optimal conditions, virtually all of the recipient mouse Ltk- Aprt- cells take up Ca Pi-DNA complexes, as determined by fluorescent dyes specific for DNA (4'',6-diaminilo-2-phenylindol dihydrochloride) or for calcium salts (chlorotetracycline). Only a small proportion of the cells have detectable Ca Pi-DNA complexes in the nucleus. Uptake of the Ca Pi-DNA complexes was highly dependent upon the pH at which the Ca Pi-DNA complex was formed and upon the concentration of DNA in the complex.