Transformation of rat embryo fibroblasts by cloned polyoma virus DNA fragments containing only part of the early region.
Transformation of rat embryo fibroblasts by cloned polyoma virus DNA fragments containing only part of the early region.
复制标题
仅含有部分早期区域的克隆多瘤病毒 DNA 片段对大鼠胚胎成纤维细胞的转化。
DOI：
10.1073/pnas.77.7.3978
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发表时间：
1980
影响因子：
11.1
通讯作者：
Moreau,PE
中科院分区：
文献类型：
--
作者：
Hassell,JA;Topp,WC;Rifkin,DB;Moreau,PE
在大肠杆菌X1776中构建并克隆含有多瘤病毒DNA的完整基因组或两个HindIII片段中的一个或另一个的重组质粒，并单独测试它们的DNA转化已建立的大鼠细胞系的能力。含有完整多瘤基因组的重组质粒和含有多瘤DNA的HindIII-1片段（45-1.4图单位）的重组质粒有效地转化大鼠细胞，而含有HindIII-2片段（1.4-45.0图单位）的质粒则不能。通过用克隆的HindIII-1片段感染建立的许多独立的转化细胞系的性质被确定。与亲本细胞系相反，用克隆的HindIII-1片段转化的大鼠细胞生长到高饱和密度，在稀释的琼脂悬浮液中高效地形成菌落，产生高水平的纤溶酶原激活剂，并显示肌动蛋白缆的无序排列。根据检查的所有标准，这些由片段转化的细胞与由完整多瘤病毒DNA转化的细胞难以区分。从许多HindIII-1片段转化的细胞系制备的细胞DNA通过Southern印迹杂交分析多瘤病毒序列的存在和排列。在所有检查的病例中，仅检测到多瘤DNA的HindIII-1片段内所含的病毒序列。这些数据建立了多瘤DNA序列映射在早期区域的有限部分和转化表型的诱导和维持之间的强相关性。
Recombinant plasmids containing either the entire polyoma viral genome or one or the other of the two HindIII fragments of polyoma virus DNA were constructed and cloned in Escherichia coli X1776, and their DNAs were individually tested for the capacity to transform an established line of rat cells. The recombinant plasmids containing the entire polyoma genome and those containing the HindIII-1 fragment of polyoma DNA (45-1.4 map units) efficiently transform rat cells, whereas the plasmids containing the HindIII-2 fragment (1.4-45.0 map units) do not. The properties of many independent transformed cell lines established by infection with the cloned HindIII-1 fragment were determined. In contrast to the parent cell line, rat cells transformed with the cloned HindIII-1 fragment grow to high saturation densities, form colonies with high efficiency in dilute agar suspension, produce high levels of plasminogen activator, and display a disorganized arrangement of actin cables. By all criteria examined, these cells transformed by fragments are indistinguishable from cells transformed by whole polyoma viral DNA. Cellular DNA prepared from many HindIII-1 fragment-transformed cell lines was analyzed for the presence and arrangement of polyoma viral sequences by Southern blot-hybridization. In all cases examined, only those viral sequences contained within the HindIII-1 fragment of polyoma DNA were detected. These data establish a strong correlation between polyoma DNA sequences mapping within a restricted portion of the early region and the induction and maintenance of the transformed phenotype.