Elongation Blockers
Elongation blockers
Elongation blockers in oligonucleotide synthesis, such as 3′-phosphate groups or dideoxynucleotides, prevent chain extension beyond a specific point. These blockers provide precise control over sequence length and enable the creation of functional structures such as molecular beacons, primers, and probes. In assays using HyProbes or LightCycler probes, elongation blockers inhibit unwanted extension, reducing non-specific amplification and increasing target specificity.
Modifications
Structure Change
Benefits
Phosphate (Phos)
Removes the 3′-OH group required for chain elongation, preventing further nucleotide addition by DNA polymerases
A simple and effective modification that retains the natural phosphodiester backbone structure.
Spacer(C3)
It consists of a three-carbon chain that replaces the 3′-OH group, making it impossible for polymerases to add nucleotides
It effectively blocks polymerase extension and can prevent exonuclease-dependent oligo cleavage.
Reverse dT (rev-dT)
Attaches a thymidine nucleotide in reverse orientation (3′-3′ linkage)
It is an effective polymerase extension blocker while preserves the ability to form base pairs at the 3′-end.
Reverse dSpacer (rev-dSpacer)
Introduces a stable abasic site at the 3′-end. It is similar to the reverse dT but lacks a nucleobase.
It introduces a stable abasic site at the 3′-end of the oligonucleotide, effectively blocking polymerase extension while minimizing potential base-pairing interactions.
Dideoxycytidine (ddC)
A synthetic analog of deoxycytidine lacking both 2′- and 3′-OH groups on the ribose.
When incorporated at the 3′-end of an oligonucleotide, it effectively blocks polymerase extension
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