Minor Groove Binder Probes

A recent improvement on classical TaqMan probes is the development of minor groove binders (MGB) chemistry.1-12-1 Modification of an oligonucleotide with an MGB such as dihydroclyclopyrroloindole tripeptide (DPI3) results in a substantial increase in the interaction of the oligonucleotide with a complementary DNA strand. The MGB effectively acts as a clamp that secures the base-pairing interaction between the oligonucleotide and the target DNA sequence (Fig. 2). The MGB interacts with the minor groove of the double-stranded DNA mainly through van der Waals forces, stretched over approximately 5 base pairs. The increase in stability is such that the binding energy of a 12-mer probe modified with an MGB group at either end is similar to the binding energy of a perfectly matching 27-mer, nonmodified oligonucleotide. For TaqMan probes, the MGB is added at the 3'end, which de facto makes the oligonucleotide nonextensible. The quencher dye can be conjugated directly to the MGB.

Unlike the TaqMan probe, where optimal discrimination of polymorphic sites is achieved by positioning the mismatch near the center of the probe, MGB-modified TaqMan probes discriminate between alleles better when the mismatch is in the 3' end of the sequence (see asterisks in Fig. 2). Base mismatches in the MGB binding region of the TaqMan probe greatly affect the ability of the MGB to bind and stabilize the base-pairing interaction, resulting in destabilization of the DNA duplex. Whereas a mismatch in the non-MGB binding region results in a decrease of 3-9°C in the melting temperature of the duplex, a mismatch in the MGB portion of the probe can result in decreases as much as 16°C.[12] Combined with the fact that MGB probes are much shorter than regular TaqMan probes (12-16 vs. 20-30 bp), this results in a superior mismatch discrimination overall when using MGB-modified probes.

Getting Started With Dumbbells

Getting Started With Dumbbells

The use of dumbbells gives you a much more comprehensive strengthening effect because the workout engages your stabilizer muscles, in addition to the muscle you may be pin-pointing. Without all of the belts and artificial stabilizers of a machine, you also engage your core muscles, which are your body's natural stabilizers.

Get My Free Ebook


Post a comment