title: Enzymatic synthesis of oligonucleotides of defined sequence. Addition of short blocks of nucleotide residues to oligonucleotide primers.
abstract: Polynucleotide phosphorylase from Escherichia coli can be used to catalyse the addition of short tracts of deoxyadenylate residues to the 3'-termini of deoxyribooligonucleotides of the type pdAn-dN (where dN = dC, dT or dG) using dADP as donor. Similarly, the enzyme can also be used to catalyse the addition of short tracts of adenylate residues to the 3'-termini of ribooligonucleotides of the type An-N (where N = C, U or G) using ADP as donor. In the ribooligonucleotide series, phosphorolytic cleavage of the primer oligonucleotides is significant and results in the concommitant production of oligoadenylates lacking the N residue. Oligomers of the same length, with and without the residue N, were readily separated by thermal elution from cellulose-pdT9 columns. This latter procedure therefore provides a simple method for purification of the oligoadenylates containing an internal base substitution and it also provides a convenient assay for oligonucleotide phosphorolysis.
mesh_terms: D000227:Adenine Nucleotides; D001483:Base Sequence; D003854:Deoxyribonucleotides / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology*; D009841:Oligonucleotides / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D010727:Phosphoric Diester Hydrolases; D011063:Poly A-U; D011119:Polynucleotides / Q000032:analysis; D011117:Polyribonucleotide Nucleotidyltransferase / Q000378:metabolism*; D006163:Ribonuclease T1; D012265:Ribonucleotides / Q000032:analysis; D012910:Snake Venoms; D013154:Spleen / Q000201:enzymology
chemical_list: D000227:Adenine Nucleotides; D003854:Deoxyribonucleotides; D009841:Oligonucleotides; D011119:Polynucleotides; D012265:Ribonucleotides; D012910:Snake Venoms; D011063:Poly A-U; D011117:Polyribonucleotide Nucleotidyltransferase; D006163:Ribonuclease T1; D010727:Phosphoric Diester Hydrolases