Real-time fluorescence quantitative PCR method for detecting NPM1 genic mutation

Abstract

The invention provides a method and a kit for detecting nucleophosmin (NPM1) genic mutation by using a primer specificity fluorescence quantitative polymerase chain reaction (PCR), and in particular relates to diagnosis of the NPM1 genic mutation in a tumor tissue and the peripheral blood serum of a tumor patient. In the kit, a specific oligonucleotide primer sequence and a probe sequence are designed for NPM1 gene 12 exon mutation, namely NPM1-mutation A, NPM1-mutation B, NPM1-mutation D and NPM1 wild type gene, fluorescence quantitative PCR detection is performed on each sample to be detected by a wild type PCR system and a mutation type PCR system respectively, and whether NPM1 gene 12 exon mutation, namely the NPM1-mutation A, the NPM1-mutation B or the NPM1-mutation D exists in the sample is judged by Ct value differences between a mutation type PCR and a wild type PCR. By the method and the kit, NPM1 gene mutation can be detected quickly and qualitatively or quantitatively, and reference basis is provided for the determination of diagnosis and treatment schemes and prognosis determination of acute myelocytic leukemia (AML) and myelodysplastic syndrome (MDS).

Description

[0001] technical field [0002] The invention relates to the field of gene mutation detection, in particular to primers, probes and methods for real-time fluorescent quantitative PCR detection of NPM1 gene mutation. Background technique [0003] The nucleophosmin (NPM) gene is located on human chromosome 5q35 and contains 12 exons. Three mRNAs and corresponding protein products were formed by different cleavage of RNA, among which the longest mRNA (NM_002520) produced a protein product of 294 amino acids, which was the most important type of NPM1. Mutations in the NPM1 gene are detected in 25% to 35% of all acute myeloid leukemia (AML) cases and in 45% to 64% of normal karyotype acute myeloid leukemia (AML-NK) , also found in myeloproliferative disorders and myeloid dysplasia. The vast majority of NPM1 gene mutations occur in exon 12, of which the most common NPM1-mutationA accounts for 75%-80% of NPM1 gene mutations, followed by NPM1-mutationB, accounting for 10% of NPM1 ...

AI Technical Summary

Problems solved by technology

Post-PCR sequencing and post-PCR dHPLC require post-PCR post-processing, which is cumbersome, time-consuming, and prone to contamination

The FISH hybridization method has many steps, takes a long time, and the detection cost is high

Detection of gene mutations in leukemia cells Due to the co-existence of mutant DNA of cancer cells and wild-type DNA of a large number of normal cells in peripheral blood, it requires high sensitivity and high specificity, and the detection ability of general sequencing and other detection methods is difficult. fulfil requirements

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