Synthesis of oligodeoxyribonucleotides containing degenerate bases and their use as primers in the polymerase chain reaction.
Heptadecaoligodeoxyribonucleotides containing one or more of the bases, 6H,8H-3,4-dihydropyrimido[4,5-c][1,2]oxazin-7-one (P), 2-amino-6-methoxyaminopurine (K), and hypoxanthine (I) and combinations of P with K and I have been synthesised on a DNA synthesiser. The stability of duplexes containing these basemodified oligomers with P/A, P/G, K/C and K/T; P/A, P/G, I/C, I/T and I/A, I/G, I/C, I/T base pairs were compared by measuring their melting transition (Tm) values. Oligomers containing both P and K and P and I were more stable than those with I alone or with mismatches. These oligomers together with one with a P base at the 3'-end were used as primers in polymerase chain reaction (PCR) experiments. They were all effective primers except one with I alone and a triple mismatch. Thus the use of the degenerate bases P and K in primer design is established.
Lin
P K
PK
MRC Laboratory of Molecular Biology, Cambridge, UK.
Brown
D M
DM
D000577:Amides; D001483:Base Sequence; D004247:DNA / Q000138:chemical synthesis*; D006860:Hydrogen Bonding; D007202:Indicators and Reagents; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis*; D010756:Phosphoric Acids; D016133:Polymerase Chain Reaction / Q000379:methods*; D013329:Structure-Activity Relationship; D013816:Thermodynamics
D000577:Amides; D007202:Indicators and Reagents; D009838:Oligodeoxyribonucleotides; D010756:Phosphoric Acids; D004247:DNA; C011067:phosphoramidic acid