Digital PCR concentration calculation method

A calculation method and concentration technology, applied in the direction of calculation, electrical digital data processing, special data processing applications, etc., can solve problems such as measurement accuracy cannot be guaranteed, and achieve the effect of broadening the concentration detection range and improving accuracy

Active Publication Date: 2018-01-23
领航医学科技(深圳)有限公司
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Problems solved by technology

[0003] Quantitative measurement of gene concentration by digital PCR based on Poisson distribution can have very high accuracy, but the measurement accuracy cannot be guaranteed for the dilution in the dynamic range of unknown samples

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Embodiment Construction

[0019] Based on the real-time cluster concentration calculation method, multi-reaction point detection is adopted, and the reaction points are monitored in real time, and data detection is performed in each reaction cycle. The data expression of the detection reaction point can be some physical or chemical quantities that can be quantified, such as light intensity, number of molecules, number of nucleic acids, number of proteins, etc. with the number of expressed molecules or single nucleic acid or protein. This detection is a dynamic detection, and the reaction point data detection is carried out from the beginning of the reaction to the end of the reaction. The multi-cycle detection data of each reaction point should be stored correspondingly, and then draw the amplification curve diagram of the reaction point after the reaction amplification is completed. Figure 1 Generally, the fluorescence amplification curve is mainly used.

[0020] When the difference in the number of...

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Abstract

The invention discloses a digital PCR concentration calculation method, which determines and calculates the initial concentration of a reactant according to the amplification cycleCt value of the real-time fluorescence quantitative curve in the PCR amplification process combined with the real-time clustering method. The fluorescence threshold R11 has an intersection with the amplification curve ofeach positive reaction unit during exponential growth, and the intersection corresponds to the corresponding amplification cycle value Cti. The digital PCR concentration calculation method specifically includes the following steps: clustering according to the Cti, obtaining k clusters, wherein the central value of each cluster is M1, M2, ..., Mk in descending order; calculating that the number ofthe Cti contained in each cluster is Sj, calculating the number of target genes in each cluster by an Mj and the Sj, adding up the number of target genes in each cluster, finally obtaining the concentration value of the initial target genes.

Description

technical field [0001] The invention relates to a digital PCR concentration calculation method. Background technique [0002] Digital PCR is to evenly distribute the fluorescence quantitative reaction system into a large number of tiny reaction units, each of which does not contain or contains one or more target gene fragments. After the amplification, the fragments containing the target gene generate a positive detection signal, and those that do not contain the target gene do not generate a detection signal. The ratio of the number of positive reaction units determined by the end-point fluorescence signal to the total reaction units is calculated by statistical methods. The copy number of the target gene in the original sample. [0003] The quantitative measurement of gene concentration by digital PCR based on Poisson distribution can have very high accuracy, but cannot guarantee the measurement accuracy under the dilution degree in the dynamic range of unknown samples. ...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G06F19/24
Inventor 程标
Owner 领航医学科技(深圳)有限公司
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