Multi-PCR primer for detecting EGFR/KRAS/BRAF genetic mutation locus based on next-generation sequencing technology and method and application

Abstract

The invention provides a multi-PCR primer for detecting EGFR / KRAS / BRAF genetic mutation locus based on a next-generation sequencing technology. The multi-PCR primer comprises two or more pairs of 14 pairs of primers shown as SEQ ID NO: 1-SEQ ID NO: 28; preferably, the multi-PCR primer comprises a primer pair including any two or three genes of amplification EGFR, KRAS and BRAF genes at a minimum, wherein each gene amplifies one or more exon regions, and one or more primer pairs are adopted for each exon region. The multi-PCR primer set can be used for efficient detection of diversity of EGFR / KRAS / BRAF genetic mutation locus and can cover 18th-21th EGFR exon, the second KRAS exon and the 15th BRAF exon.

Description

Technical field [0001] The invention belongs to the field of molecular biology, and particularly relates to a multiple PCR primer based on next-generation sequencing technology for detecting EGFR / KRAS / BRAF gene mutation sites, a method and application. Background technique [0002] Epidermal growth factor receptor (EGFR) is the expression product of the proto-oncogene C-erbB-1 (HER-1) located on the short arm of chromosome 7. It is the 4 epidermal growth factor receptor family (HER) One of the members, the HER family plays an important regulatory role in cell physiological processes. The mutations in the EGFR tyrosine kinase region mainly occur in exons 18-21, among which exon 19 and 21 mutations account for 90% of the total mutations. Abnormal EGFR signal transduction is the cause of many kinds of tumors. Studies have shown that EGFR is expressed in a variety of tumors, such as colorectal cancer, breast cancer, pancreatic cancer, prostate cancer and non-small cell lung cancer....

AI Technical Summary

Problems solved by technology

In the Sanger sequencing method, a single pair of primers can detect multiple mutations, but the mutation sites on multiple genes or different exons of the same gene need to be amplified and sequenced separately, which is cumbersome and has a low sensitivity of about 20 %, higher false negative rate

Although the fluorescent quantitative PCR method has high sensitivity, each pair of primers can only detect one mutation, and each mutation requires a separate PCR reaction system, and the simultaneous detection of multiple mutation sites in multiple samples is cumbersome.

These two methods require a large amount of samples and are not suitable for detecting multiple gene mutation sites at the same time

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