Chu B C, Orgel L E
Salk Institute for Biological Studies, San Diego, CA 92138.
DNA Cell Biol. 1990 Jan-Feb;9(1):71-6. doi: 10.1089/dna.1990.9.71.
A simple, efficient procedure for cross-linking two complementary oligonucleotides, which does not require chemical modification of either oligonucleotide, is described. One of the oligonucleotides is first converted to the 5'-phosphorothioate derivative with polynucleotide kinase. It is then incubated with its complement in the presence of 1 microM trans-platinum(II)diammine dichloride. After overnight incubation, 40-50% cross-linking is observed. DNA synthesis by the Klenow fragment of Escherichia coli DNA polymerase I is blocked at the cross-linked site, resulting in the formation of truncated products. Potassium platinous chloride (K2PtCl4) and cis-platinum(II)diammine dichloride form cross-links less efficiently than the trans isomer.
本文描述了一种简单、高效的交联两个互补寡核苷酸的方法，该方法不需要对任何一个寡核苷酸进行化学修饰。首先用多核苷酸激酶将其中一个寡核苷酸转化为5'-硫代磷酸酯衍生物。然后在1微摩尔反式二氯二氨合铂（II）存在下与它的互补链一起温育。过夜温育后，观察到40-50%的交联。大肠杆菌DNA聚合酶I的Klenow片段的DNA合成在交联位点被阻断，导致形成截短产物。氯化亚铂酸钾（K2PtCl4）和顺式二氯二氨合铂（II）形成交联的效率低于反式异构体。
