|
Many molecular biology applications require oligonucleotide primers and/or probes with enhanced duplex stability, that is, a higher affinity for their complementary sequences. Examples include antisense, siRNA, and SNP detection experiments. For these and other applications, enhancing primer/probe-target duplex stability increases both the selectivity and sensitivity of the oligo for its target. An oligo’s binding affinity for its target can be manipulated by incorporating various modifications into it, either on the ends or at an internal position (1). Examples include 2’-OMethyl base, 2-Amino-dA, 5-methyl-dC and 2’-Fluoro bases. Changes in duplex stability incurring from such modifications is indicated by changes in the melting temperature (Tm) of the duplex formed between the modified oligo and its target over that formed by the unmodified oligo. Depending on the modification, Tm can be increased over a range of 0.5C to 3.0C per base substitution.
|
