3'-AMINO-MODIFIED NUCLEOTIDES USEFUL AS POTENT CHAIN TERMINATORS FOR CURRENT DNA-SEQUENCING METHODS

We describe the synthesis of modified nucleoside triphosphates of the four DNA bases containing a 3'-amino group which were prepared from th e corresponding 3'-azido derivatives. Introduction of the triphosphate and subsequent reduction of the N3 to the NH2 group led directly to t he target molecules 6a-d. Furthermore, 3'-amino-2',3'-dideoxynucleosid e 5'-triphosphates proved to be potent inhibitors of the enzymatic syn thesis of DNA catalyzed by the standard sequencing enzymes T7 DNA poly merase, sequenase version 2.0, Thermus aquaticus DNA polymerase, and T hermus thermophilus DNA polymerase. Both radioactive and fluorescent s equencing methods were applied successfully to the 3'-amino-modified t erminators. Investigations in view of using these chain terminators ac cording to Sanger's sequencing method for fluorescence labeling were d one.