A recent development has been the use of probes incorporating peptide nucleic acid (PNA) or locked nucleic acid (LNA™). In PNA the entire deoxyribose phosphate backbone is replaced by a structurally homo-morphous backbone consisting of repeating N-(2-amino-ethyl)-glycine units linked by peptide bonds. As the PNA backbone is not charged, hybridization is not affected by intrastrand repulsion, and no salt is necessary to facilitate and stabilize the formation of PNA and DNA or RNA duplexes. Furthermore, the DTm of a single PNA/DNA mismatch is significantly higher than that of a DNA/DNA mismatch, making PNA probes useful for mismatch detection. PNA technology has been used in an ingenious approach utilizing quencher-labeled primers. This system uses two labeled molecules: 1) a quencher-labeled PNA probe with a C-terminal DABCYL group; and 2) a primer with a target-specific sequence at its 3'-end and a fluorophore and a PNA-probe-complementary sequence tag at its 5'-end. The PNA/primer duplex has a Tm higher than the primer annealing temperature, but lower than the Tm of the primer/amplicon duplex. This ensures that excess primer is quenched at the annealing temperature and the fluorescence measured during the annealing step indicates the amount of primer hybridized to amplicon, plus any full-length ds amplicon as the end result is a fluorescently labeled ds amplicon. The main disadvantage with PNA probes is that they can aggregate and precipitate.
In LNA the furanose ring conformation is restricted by a methylene bridge that connects the 2'-oxygen position of ribose to the 4'-carbon. This bridge reduces the conformational flexibility of the ribose, which imparts superior affinity and specificity in binding complementary sequences of DNA or RNA. In addition, the change in Tm caused by a mismatch is significantly greater with a LNA/DNA duplex than with of a DNA/DNA duplex, resulting in enhanced specificity for SNP/mutation analyses. Using appropriate dyes, it is possible to use probes as short as 7 nucleotides long for mismatch discrimination, making it possible to generate universal genotyping reagents. Unlike PNA, there is no problem with solubility of LNA molecules.
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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.