The resulting melting curves on most instruments show 2 melting regions, a lower temperature region of heteroduplex melting and a higher region of homoduplex
For dHPLC, it has been shown (1) that temperature conditions can be optimized by use of predictive software to improve the separation of heteroduplex and homoduplex peaks and achieve high-resolution results.
The model is extended to include the case in which the component of the mixture that is present in proportion 1 - x is heterozygous, sharing one allele with the homozygous reference DNA, so that half of this proportion contributes to each type of homoduplex initially present in the previous analysis.
It relies on the use of a hydrophobic column based on reversed-phase liquid chromatography for the separation of heteroduplex and homoduplex DNA at specific optimized temperatures and has recently been used by several research groups to study mitochondrial DNA (mtDNA) mutation (1-6).
We found the optimal temperature (the temperature that results in the best resolution of the heteroduplex and homoduplex peak) to be 59 [degrees]C (data not shown).
DHPLC separates heteroduplex and homoduplex
molecules by ion-pair chromatography under partially denaturing conditions.
The continuing development of instruments with finer temperature control and fluorescence acquisition will lead to increased detail derived from heteroduplex and homoduplex
contributions to the overall melting curve, providing even greater ability to identify unique sequence variants.
The resulting melting curve is a composite of both heteroduplex and homoduplex
The more stable homoduplex
polymorphism sample (GC base pair) was distinguished by a higher [T.
With the melting temperature predicted by the Navigator software, we were able to distinguish heteroduplex from homoduplex
Our approach is based on 3 principles: (a) a single-base mismatch produces a local conformational change in the double-stranded DNA, leading to differential migration of the heteroduplex and homoduplex
bands; (b) the addition to the gel of [Zn.
The 3 heterozygotes are all in the same SNP class (12), giving the same heteroduplex mismatches (C:A and T:G) and homoduplex
matches (C:G and A:T).