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Denaturation mapping
Denaturation Mapping is a form of optical mapping, first described in 1966. It is used to characterize DNA molecules without the need for amplification or sequencing. It is based on the differences between the melting temperatures of AT-rich and GC-rich regions.〔Inman RB. (1966)."A denaturation map of the lamda phage DNA molecule determined by electron microscopy". Journal of Molecular Biology 18: pp. 464-476.〕 Even though modern sequencing methods reduced the need for denaturation mapping, it is still being used for specific purposes, such as detection of large scale structural variants.〔Reisner W;Larsen N B; Silahtaroglu A; Kristensen A; Tommerup N; Tegenfeldt J O; Flyvbjerg H. (2010). "Single-molecule denaturation mapping of DNA in nanofluidic channels". PNAS. doi: 10.1073〕
==Methodology==
When subjected to denaturing factors like increased heat or chemicals like formamide in low levels, DNA is partially denatured in a predictable pattern based on its nucleotide content in different regions.〔 This allows unique fingerprints or ‘barcodes' to be generated for molecules with different sequences not unlike restriction mapping. In the earliest forms of denaturation mapping, DNA was denatured by heating in presence of formaldehyde 〔 or glyoxal〔Johnson D. (1975)."A new method of DNA denaturation mapping". Nucleid Acids Research 2(11): pp. 2049-2053.〕 and visualized using electron microscopy. Dyes that selectively bind to double stranded DNA like ethidium bromide could be used to monitor the extend of denaturation. But it was not possible to observe locations of denaturation based on this information. And requirement of electron microscopy made this method more strenuous to perform.
More recently microfluidics were used for denaturation mapping of single molecules. In this method a reservoir that contains individual stained DNA molecules is placed next to nanochannels. Under pressure the molecules are pushed towards the nanochannels, and stretch out. With the application of heat in the presence of formamide the molecules denature and while in these channels, denaturation profile of these molecules can be observed by fluorescence microscopy.〔
抄文引用元・出典: フリー百科事典『 ウィキペディア（Wikipedia）』
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