Primers, reagent kit and method for detecting IDH1 (isocitrate dehydrogenase 1) R132H gene variation by aid of ddPCR (droplet digital polymerase chain reaction) technologies

Abstract

The invention discloses primers, a reagent kit and a method for detecting IDH1 (isocitrate dehydrogenase 1) R132H gene variation by the aid of ddPCR (droplet digital polymerase chain reaction) technologies. The method includes extracting cfDNA (cell-free deoxyribonucleic acid) in peripheral blood; designing two probes and a pair of primers for IDH1R132H mutant types and wild types; carrying out ddPCR amplification by the aid of the cfDNA used as a template; carrying out data analysis on obtained amplification products to obtain absolute quantities and proportions of IDH1R132H mutant genes. Thereagent kit for detecting the IDH1R132H gene variation is based on the method. The primers, the reagent kit and the method have the advantages that provided primer amplification fragments 83bp are particularly applicable to amplifying small-fragment DNA samples such as plasma free DNA and can be combined with the detection probes, accordingly, the IDH1R132H gene variation with extremely low abundance can be detected from the cfDNA, the primers, the reagent kit and the method are good in specificity and high in sensitivity, human IDH1R132H gene variation can be absolutely quantitatively detected by the aid of the primers, the reagent kit and the method, and important effects can be realized by the primers, the reagent kit and the method for guiding clinical treatment and improving patientprognosis; the IDH1R132H gene variation can be detected only by the aid of the peripheral blood without tissue specimen collection, and accordingly the method is particularly suitable for patients intolerant to tissue sampling.

Description

technical field [0001] The invention relates to the fields of medicine and biotechnology, in particular to a primer, a kit and a detection method for detecting IDH1 R132H gene variation by ddPCR technology. Background technique [0002] Glioma is the most common primary central nervous system tumor, accounting for about 35% to 60% of all primary intracranial tumors, with an adult incidence of 8 to 10 / 100,000 and a 5-year survival rate of 20% to 30%, and its incidence rate is still on the rise in recent years. The World Health Organization (WHO) has established clinical guidelines for classifying gliomas into various subtypes and grading them from I to IV indicating their degree of malignancy. [0003] Isocitrate dehydrogenase (IDH) is an enzyme family that plays an important role in the tricarboxylic acid cycle, not only plays an important role in energy metabolism, amino acid and vitamin synthesis, but also regulates the activity of the enzyme will Directly affect IDH or ...

AI Technical Summary

Problems solved by technology

However, due to the extremely low content of plasma cell-free DNA in the blood and the interference of a large amount of blood-derived DNA, it is still difficult to separate and purify DNA with high-efficiency DNA enrichment methods and high-sensitivity detection methods (such as sequencing, ARMS-PCR). Accurate and reliable detection of specific species and sequences of tumor-associated cell-free DNA

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