Method for preparing gene mutation reference substance

Abstract

The invention discloses a method for preparing a gene mutation reference substance. The method disclosed by the invention comprises the following steps: firstly, mixing mutant genome DNA and wild genome DNA according to at least two proportions, measuring the mutation proportion of the prepared reference substance, and calculating a slope and intercept of a fitting straight line; and then preparing the mutation reference substance with any mutation proportion which does not exceed the initial mutation type genome DNA mutation proportion according to the target requirement without verifying themutation proportion of the reference substance obtained by dilution.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for preparing a gene mutation reference product by utilizing mutant genome DNA and wild-type genome DNA. Background technique [0002] In the field of cancer treatment, specific mutations carried by multiple genes are closely related to the efficacy of cancer targeted therapy drugs. Therefore, gene mutation detection has important clinical significance, and gene mutation detection technology has also been widely used. In the development and verification process of gene mutation detection technology, it is necessary to use reference products containing target gene mutations for the development and performance verification of detection reagents, which are characterized by simulating clinical samples as much as possible and containing different and known mutations Proportion. [0003] When mixing genomic DNA extracted from mutant and wild-type clinical samples or c...

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Problems solved by technology

[0005] Currently, digital PCR and high-throughput sequencing are commonly used to accurately measure the proportion of target gene mutations in gene mutation reference products, and the cost of each measurement reaction is relatively high. Therefore, when preparing mutation reference products according to conventional methods, It is necessary to prepare multiple gradients and then determine the mutation ratio one by one. This process is complicated, and the number of samples for mutation ratio determination leads to a long verification process and high verification costs, which greatly restricts the preparation efficiency of mutation reference products.

If no measurement is made on the actual mutation ratio of the target gene in the diluted mutation reference products in order to save costs, the actual mutation ratio of the mutation reference products may deviate greatly from expectations, resulting in distortion of the experimental results obtained with these reference products

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