Gene mutation analysis judgment method based on real-time fluorescence quantification PCR (polymerase chain reaction) platform

Abstract

The invention discloses a gene mutation analysis judgment method based on a real-time fluorescence quantification PCR (polymerase chain reaction) platform. A fluorescent quantitation PCR amplification result of a target gene is used for analysis judgment. The analysis judgment method comprises steps as follows: step one, after a PCR experiment ends, an experimental result is subjected to CT (computerized tomography) value calculation, and a CT value is judged to determine whether the experiment condition is continuously analyzed; step two, internal control is operated, and whether a DNA (deoxyribonucleic acid) sample is qualified is judged according to the internal control result; step three, a positive quality control product is operated, and whether an experiment system is qualified is judged according to the CT value of the positive quality control product; and step four, if the CT value is qualified through check analysis, a Tm calling program is operated, and whether the sample has mutation is judged through melting curve analysis and CT value analysis. According to gene mutation analysis judgment method, a PCR result is clearly and objectively judged, sample gene types are judged according to amplification data and a melting curve, and the misjudgment rate is low.

Description

technical field [0001] The invention relates to the field of gene mutation analysis, in particular to a gene mutation analysis and determination method based on a real-time fluorescent quantitative PCR platform. Background technique [0002] At present, the existing methods for identifying genetic mutations mainly include the following: [0003] (1) High resolution melting curve (high resolution melting, HRM). The high-resolution melting curve is based on the physical properties of nucleic acid, through the binding of saturated dyes to PCR amplification products, and monitoring the changes in the melting curve of the product to analyze gene mutations. The detection sensitivity is about 5%, and the specific location of the positive result cannot be determined, and it needs to be confirmed by sequencing. [0004] (2) Probe amplification retardation mutation method (amplification refractory mutation system, ARMS). Using the principle that the base at the 3' end of the PCR pr...

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Problems solved by technology

[0006] The detection sensitivity of the above methods is low, subjectivity is strong, and it is easy to cause misjudgment

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