DNA synthesis in human lymphocyts: intermediates in DNA synthesis, in vitro and in vivo.
DNA synthesis in human lymphocyts: intermediates in DNA synthesis, in vitro and in vivo.
复制标题
人类淋巴细胞中的 DNA 合成：体外和体内 DNA 合成的中间体。
DOI：
10.1016/s0022-2836(75)80149-5
复制
发表时间：
1975
影响因子：
5.6
通讯作者：
M. Goulian
中科院分区：
文献类型：
--
作者：
B. Tseng;M. Goulian
两种体外系统，一种使用全细胞裂解液，另一种使用从人淋巴细胞系制备的洗涤细胞核悬浮液，其特征在于它们对DNA合成的要求和合成产物的性质。三磷酸核糖核苷对DNA合成有刺激作用，这不能用它们维持脱氧核糖核苷三磷酸水平的作用来解释。裂解物的合成速率约为其硼酸盐的10%，并在20至40分钟内呈线性；在那些活跃的细胞核中，最多有1%的细胞DNA是新合成的。一类片段，平均链长为120个核苷酸，可以在聚丙烯酰胺凝胶上区分，是整个细胞裂解物中最早标记的中间体。这些短片段长到几百个核苷酸，然后附着在高分子量的DNA上。他们的体外合成产品由数百万道尔顿大小的连续链组成。它们在体外的移动速度，也可能是单个链的生长速度，大约是相应体内生物的10%。洗涤后的细胞核产生与裂解物相似的120个核苷酸的中间产物；然而，细胞核不合成高分子量的DNA，最大的产物约为10 s。如果将细胞质提取物添加到细胞核中，则可以恢复一些合成大分子量DNA的能力。在用放射性胸腺嘧啶标记的完整的指数生长细胞中，发现了一个类似的120个核苷酸的DNA片段。
Twoin vitrosystems, one using a whole cell lysate, and the other a suspension of washed nuclei prepared from a human lymphocyte line, are characterized with respect to their requirements for the synthesis of DNA and to the nature of the synthetic product.Ribonucleoside triphosphates have a stimulatory effect on DNA synthesis that cannot be accounted for by their effect in maintaining deoxyribonucleosidetriphosphate levels. The rate of synthesis in the lysate is approximately 10% of thein vivorate and is linear for 20 to 40 minutes; in those nuclei that are active, a maximum of 1% of the cellular DNA is newly synthesized.A class of fragments, with a mean chain length of 120 nucleotides, can be distinguished on polyacrylamide gels and is the earliest labeled intermediate in the whole cell lysate. These short fragments grow to several hundred nucleotides and then attach to high molecular weight DNA. Thein vitrosynthetic product consists of continuous strands several million daltons in size. Thein vitrofork movement rate, and probably also the individual chain growth rate, are approximately 10% of the correspondingin vivorates.Washed nuclei make 120-nucleotide intermediates similar to those of the lysate; nuclei, however, do not synthesize high molecular weight DNA, the largest product being around 10 S. If cytoplasmic extract is added back to nuclei some of the ability to synthesize large DNA is restored.In intact exponentially growing cells labeled with radioactive thymidine, a similar 120-nucleotide DNA fragment has been identified.