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Determination of melting temperatures by equation-less methods

A technology of melting temperature and melting curve, applied in the field of determining one or more melting temperatures of oligonucleotide samples, which can solve the problem of no beneficial effect of treatment

Active Publication Date: 2010-07-21
F HOFFMANN LA ROCHE & CO AG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Patients whose KRAS gene is wild-type benefit from the treatment, whereas patients with a mutant variant of the gene have no beneficial effect

Method used

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  • Determination of melting temperatures by equation-less methods
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  • Determination of melting temperatures by equation-less methods

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Embodiment Construction

[0051] The present invention provides systems and methods for determining the melting temperature Tm of DNA.

[0052] The present invention provides systems and methods for determining melting temperatures by analyzing data representing melting curves. In certain aspects, a numerical determination of the first derivative of the melting curve data set is performed. A Gaussian mixture model (GMM) function with parameters determined using the Levenberg-Marquardt (LM) regression procedure was used to find an approximation to the first derivative curve. The numerically determined maximum value of the first derivative value is used as the initial condition for the parameters of the GMM function. The determined parameters provide fractional melting temperature Tm values. The Tm value is then returned and may be displayed or otherwise used for further processing.

[0053] An example of a melting curve in the context of a PCR process such as figure 1 shown. Such as figure 1 As sh...

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Abstract

Numerical determinations of the first derivatives of a melt curve data set are made. A baseline is determined for the first derivative values and the baseline is subtracted from the first derivative values to produce modified first derivative values. A first maximum value of the modified first derivative values is determined and said first maximum value represents a melting temperature Tm of a DNA sample. A model function, such as a Gaussian Mixture Model (GMM) function, with parameters determined using a Levenberg-Marquardt (LM) regression process can also be used to find an approximation to the first derivative curve. The maximum values of the numerically determined first derivative values are used as initial conditions for parameters of the model function. The determined parameters provide one or more fractional melting temperature values, which can be returned, for example, displayed or otherwise used for further processing.

Description

technical field [0001] In general, the present invention relates to processing data indicative of melting properties of oligonucleotides, and more particularly to systems and methods for determining one or more melting temperatures of an oligonucleotide sample from melting curve data. Background technique [0002] Determination of DNA melting temperature, usually performed directly after PCR experiments, is an important method for distinguishing genotypes. For example, it has been recently discussed in the literature that examination of the KRAS gene is used to determine which patients may be candidates for treatment of non-small cell lung cancer. Patients whose KRAS gene was wild-type benefited from the treatment, whereas patients with a mutant variant of the gene had no beneficial effect. Since these treatments often have major side effects, it is very important to determine the correct genotype of the patient. The use of the fused KRAS assay can help differentiate patie...

Claims

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Application Information

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IPC IPC(8): G06F19/00C12M1/40
CPCC12Q1/6813G06F17/18C12Q2527/107C12Q2537/165
Inventor R·T·克尼克
Owner F HOFFMANN LA ROCHE & CO AG