A simple procedure for cross-linking complementary oligonucleotides.
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.
Chu
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BC
Salk Institute for Biological Studies, San Diego, CA 92138.
Orgel
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LE
D001483:Base Sequence; D002945:Cisplatin; D003432:Cross-Linking Reagents; D005821:Genetic Techniques; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D009838:Oligodeoxyribonucleotides; D009841:Oligonucleotides; D011116:Polynucleotide 5'-Hydroxyl-Kinase; D013873:Thionucleotides
D003432:Cross-Linking Reagents; D009838:Oligodeoxyribonucleotides; D009841:Oligonucleotides; D013873:Thionucleotides; C034697:transplatin; D011116:Polynucleotide 5'-Hydroxyl-Kinase; D002945:Cisplatin