| Abstract: | Sanger method was modified to fulfill the requirements of sequencing of oligodeoxyribonucleotides. E. coli DNA polymerase I Klenow fragment was used for all the reactions. The method consists of three steps made in succession in one tube: 1. Optional hydrolysis of a 5'-labeled oligodeoxyribonucleotide primer in order to get a set of primers of different lengths. 2. Elongation of the produced set of primers in the presence of a template, natural dNTPs and chain terminating dNTP analogs. 3. Hydrolysis of the products of the previous step in order to remove the unterminated molecules. Change of steps in achieved just by varying the reaction conditions without any product purification. The method in insensitive to the presence of admixture of oligonucleotides which is not complementary to the primer or to the template. |
