Extraction-free reagent for fluorescent PCR of blood samples and its application

Abstract

An extraction-free reagent for fluorescent PCR of blood samples, specifically comprising: blood dilution liquid and PCR reaction liquid, the extraction-free reagent and blood sample form a PCR reaction system, wherein the alcohol substance in the blood dilution liquid is PEG200, alkaline The substance is NaOH; and the concentration of each substance in the blood dilution solution is: PEG200 content is 0.01wt% ~ 0.1wt%, NaOH content is 10mM ~ 100mM, Tris-HCl content is 5mM ~ 50mM, Tween type The content of the nonionic surfactant is 0.1wt%-1wt%. The extraction-free reagent in the present invention can obtain unexpected system stability and reduce the interference ability of inhibitors in the blood, which improves the accuracy of the test results. The extraction-free reagent can fully denature various inhibitors in the blood, and subsequent PCR The system's tolerance to inhibitors has been effectively improved.

Description

technical field [0001] The invention belongs to the field of molecular biology. Specifically, the invention relates to an extraction-free reagent and an application method for direct fluorescent PCR amplification of blood suitable for Taq man SNP typing. Background technique [0002] In recent years, fluorescent PCR technology has been more and more widely used in clinical and inspection and quarantine fields due to its fast, accurate and simple characteristics. Taq man SNP typing technology is one of the important applications of fluorescent PCR technology. It is widely used in the detection of pathogen drug resistance mutations, genetic disease detection, prenatal and postnatal care, and tumor-targeted drug target detection. It is an important technical means and has been widely promoted in clinic. [0003] Human genomic DNA is an important analytical sample for clinical Taq man SNP typing detection, and blood is an important biological source commonly used in clinical pr...

AI Technical Summary

Problems solved by technology

[0005] In addition, the prior art CN109402239A discloses an extraction-free direct amplification reagent for real-time fluorescent quantitative PCR, which is used for the detection of nucleic acids in non-blood samples such as respiratory secretions, but the types of inhibitory substances in different types of samples in this field are different The reagents used in the prior art are used in DNA in the blood for Taq man SNP typing detection, and the result has a problem of low accuracy, which may be due to the high content and amount of interfering substances in the blood, resulting in the inhibition of substances Not easy to effectively denature

Images