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High Resolution Melt (HRM) analysis is a powerful technique in molecular biology for the detection of mutations, polymorphisms and epigenetic differences in double-stranded DNA samples. It was discovered and developed by Idaho Technology and the University of Utah. [1] It has advantages over other genotyping technologies, namely:
Except for the C/G initiation term, the first term represents the free energy of the first base pair, CG, in the absence of a nearest neighbor. The second term includes both the free energy of formation of the second base pair, GC, and stacking interaction between this base pair and the previous base pair. The remaining terms are similarly defined.
In this study they used thiazole orange as the indicator. The helicase unwinds the dsDNA to make ssDNA. The fluorescence intensity of thiazole orange has a greater affinity for dsDNA than ssDNA and its fluorescence intensity increases when it is bound to dsDNA than when it is unbound. [47] [48]
DNA nanotechnology is the field that seeks to design nanoscale structures using the molecular recognition properties of DNA molecules. [ 178 ] DNA nanotechnology uses the unique molecular recognition properties of DNA and other nucleic acids to create self-assembling branched DNA complexes with useful properties. [ 179 ]
During the rolling circle replication, the ssDNA of geminivirus is converted to dsDNA and Rep is then attached to the dsDNA at the origin sequence TAATATTAC. After Rep, along with other replication proteins, binds to the dsDNA it forms a stem loop where the DNA is then cleaved at the nanomer sequence causing a displacement of the strand.
A drawback of this method is that the product should be purified from double stranded DNA (dsDNA) and other left-over material from the PCR reaction. Enzymatic degradation of the unwanted strand can be performed by tagging this strand using a phosphate-probed primer, as it is recognized by enzymes such as Lambda exonuclease .
At a wavelength of 260 nm, the average extinction coefficient for double-stranded DNA (dsDNA) is 0.020 (μg/mL) −1 cm −1, for single-stranded DNA (ssDNA) it is 0.027 (μg/mL) −1 cm −1, for single-stranded RNA (ssRNA) it is 0.025 (μg/mL) −1 cm −1 and for short single-stranded oligonucleotides it is dependent on the length and base ...
On the left is a drawing of the complex formed between alpha-hemolysin and dsDNA with linkage through an oligomer.On the right, movement of this complex in relation to a nanopore channel is shown sequentially in two steps (I) and (II).