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Genetic assay based on hybridization with a multivalent probe, wherein the probe contains multiple sequences that are complimentary to multiple sequences on the target

IP.com Disclosure Number: IPCOM000004359D
Publication Date: 2000-Oct-13
Document File: 1 page(s) / 3K

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Related Documents

"Principles of Polymer Chemistry" by Paul J. Flory (Cornell Univ Press; ISBN: 0801401348 1953): OTHER

Abstract

The present invention is for a genetic assay based on hybridization with a multivalent probe, wherein the probe contains multiple sequences that are complimentary to multiple sequences on the target. If the target is present during the hybridization, molecular weight increase or even gelation occurs, which can be detected either via light scattering, rheological, or molecular weight measurements.

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Formation of crosslinks in polymers leads to a rapid increase in average molecular weight. Eventually, an infinite network is formed and the gelation point is reached. With polymers of intermediate to high molecular weight, this gel point can be reached relatively quickly, even with a small number of crosslinks. Quantitative treatment of the theory of gelation can be found in "Principles of Polymer Chemistry" by Paul J. Flory (Cornell Univ Press; ISBN: 0801401348 1953).

A number of genetic assays are based on a sequence of 1) denaturation of double stranded DNA molecule 2) mixing in a relatively short DNA probe (anchored to a solid support) that is complementary to that which is being assayed (often tagged with a radioactive or fluorescent moiety to facilitate analysis) 3) hybridization, during which the probe forms double stranded DNA with the target. Once hybridization occurs, the solid can be rinsed free of any un-hybridized DNA. Detection of the radioactive or fluorescent moiety on the rinsed solid support constitutes a positive test. This method is very expensive, since the preparation of probes containing radioactive or fluorescent moieties is very expensive. The process of anchoring them to the surface adds additional cost.

The present invention is for a genetic assay based on hybridization with a multivalent probe, wherein the probe contains multiple sequences that are complimentary to multiple sequences on the target. If the target is present during the hybridization, molecular weight increase or even gelation occurs, which can be detected either via light scattering, rheological, or molecular weight measurements.

Figure 1 (attached), shows a schematic of this sequence. In this example, the target analyte has the structure A-G-C-C-G-T-T-A-G-C-T-A-(X)-G-G-C-G-A-T-T-T-A-T-A-A, wherein C, T, G, and A have their conventional meanings of cytosine, thymine, guanine, and adenine, respectively. (X) represents additional segments of a DNA molecule. For purposes of the present invention, the focus is on two or more known segments (target segments) of the molecule (the two chain ends in this example). The probe is selected to have a complimentary sequence to each target segment to facilitate network formation. Hence the probe has the following structure: C-C-G-C-T-A-A-A-T-A-T-T-(Y)-T-C-G-G-C-A-A-T-C-G-A-T, wherein the group (Y) represents a connecting group, which can be any divalent group know in the art. The result of the assay, after hybridization, is a network, as outlined in Figure 1.

It is easy to imaging a tetravalent probe, wherein each probe molecule contains four arms that are complimentary to segments in the target DNA, such that each probe molecule becomes attached to four different sequences of target DNA molecules. Many other embodiments become obvious, including trivalent and pentavalent probes; tetravalent probes with two sets of equivalent sequences (e.g. (C-C-G-C-T-...