The preparative synthesis of oligodeoxyribonucleotides using RNA ligase.
The synthesis of nmol quantities of defined sequences of oligodeoxyribonucleotides using T4 RNA ligase has been demonstrated. Reacting using from 18 to 200 nmol of substrates in which a single 2'-deoxyribonucleoside 3',5'-bisphosphate was added to an oligodeoxyribonucleotide resulted in yields from 13 to 95%. When two oligodeoxyribonucleotides were similarly joined using RNA ligase, the yields ranged from 10 to 50%. Although the reactions contained high concentrations of enzyme and were incubated from 5 to 21 days, there was little degradation of either substrates or products. We have also characterized an unusual product which arises when 3'-phosphate terminated oligodeoxyribonucleotides are incubated with RNA ligase and high concentrations of ATP. This product has an adenylyl group linked to the 3'-phosphate by an anhydride bond. The mechanistic and synthetic implications of forming this product are discussed.
D001483:Base Sequence; D004926:Escherichia coli / Q000201:enzymology; D008722:Methods; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis*; D009841:Oligonucleotides / Q000138:chemical synthesis*; D010761:Phosphorus Radioisotopes; D011118:Polynucleotide Ligases / Q000378:metabolism*; D011130:RNA Ligase (ATP) / Q000378:metabolism*; D013379:Substrate Specificity; D013604:T-Phages / Q000201:enzymology
D009838:Oligodeoxyribonucleotides; D009841:Oligonucleotides; D010761:Phosphorus Radioisotopes; D011118:Polynucleotide Ligases; D011130:RNA Ligase (ATP)