Method of Solution-based Scanning for Alterations in a DNA Segment Using a Double-stranded DNA Binding Dye and Fluorescence Melting Profiles
This invention provides a new, solution-based method to determine whether a DNA sequence is identical to a wild-type sequence. The alteration may be a point mutation (such as a transversion or transition) or another mutation (such as an insertion or deletion). This technique uses a double-stranded DNA binding dye and fluorescent melting profiles to detect even a single mutation within a DNA segment.
In this invention, a guanine-cytosine (GC)-rich DNA segment is attached to a DNA segment of interest that is then labeled with a fluorescent marker and mixed with a denaturant. With the GC clamp attached, the DNA segment of interest has two melting domains: a higher domain associated with the clamp, and a lower domain associated with the labeled DNA segment of interest. The wild-type DNA sequence is compared to the clamped segment or to a heteroduplex composed of equal parts of the DNA segment of interest and the wild-type sequence. A difference in melting temperatures between the wild-type sequence and the DNA sequence (or the heteroduplex) indicates an alteration in the DNA sequence.
Many genetic disorders and some cancers are known to be caused by a point mutation, and can be hard to detect. Many of the currently available techniques used for the detection of point mutations are costly, not very sensitive and labor intensive. This invention provides a new, highly sensitive, fast and accurate method to determine whether a DNA sequence is identical to the wild-type sequence. This technique uses a double-stranded DNA binding dye and fluorescent melting profiles to detect even a single mutation within a DNA segment.
Stage of Development
U.S. patent number 6,346,386 has been issued on this technology. It is available for licensing.
Kojo S.J. Elenitoba-Johnson
Type of Offer:
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