A fluorescent PCR detection kit for human interleukin 28b gene polymorphism

Abstract

The invention relates to a kit for detecting the polymorphism of an interleukin 28B (IL28B) gene by utilizing a fluorescence PCR (Polymerase Chain Reaction) technology. The kit is characterized in that three primers are adopted in a single reaction tube to augment sequences of CC type and TT type of an rs12979860 gene, and the CC type and the TT type of the gene are detected by two MGB fluorescence probes with different wavelengths. The kit is simple in operation and accurate and reliable in detection result, can be applied to detection of the polymorphism of the IL28B rs12979860 gene of a patient with clinical hepatitis C and further predicts the treatment effect of antiviral drugs.

Description

technical field [0001] The invention relates to the field of medical molecular biology, in particular to a human interleukin 28B gene polymorphism fluorescent PCR detection kit. Background technique [0002] Human interleukin 28B gene (Interleukin 28B, IL28B) is located on the short arm of human chromosome 19 (19q13), encoding interferon λ3 (INF-λ3) and participates in human antiviral immune response. IL28B can stimulate the release of interferon and increase cytotoxic T lymphocytes. The number of cells, the current research shows that the single nucleotide polymorphism (SNP) rs12979860 near the upstream of the IL28B gene about 3kb, combined with peginterferon and ribavirin has a significant effect on the treatment of hepatitis C (Tanaka Y, et al, Nat Genet, 2009, 41:1105-1109), wherein the rs12979860 polymorphism CC type has a 2-3 times higher viral clearance rate than CT heterozygous type, TT type hepatitis C or hepatitis C, and CC type is higher than non-CC type C Patien...

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Problems solved by technology

[0003] Gene sequencing is the gold standard method for detecting IL28B rs12979860 gene polymorphism, but it has the disadvantages of high operational technical requirements, low detection sensitivity, and inability to distinguish heterozygous types. In recent years, allele-specific PCR (AS-PCR), Literature reports on the detection of IL28Brs12979860 gene polymorphism by real-time fluorescent PCR technology, melting curve or high-resolution melting curve analysis (HRM) (J Molecular Diagnostics, 2011, 13(4): 446-51; JClin Microbiol, 2012: 50: 3353-55; Chinese Journal of Laboratory Medicine, 2013, 36(1): 59-62 and 36(8): 722-26), in which AS-PCR needs to use gel electrophoresis to observe the band size judgment results, the detection sensitivity is low, The operation is inconvenient; the fluorescent PCR detection reported in the literature either uses single-wavelength separate tubes to detect different rs12979860 gene subtypes, which is complicated to operate, or uses a single tube for detection but there is a problem of cross-effects between different subtypes, while HRM analysis has higher requirements for instruments. The results of morphism detection are affected by the instrument, and the shortcomings of these detection methods limit the clinical application of IL28B rs12979860 gene polymorphism detection

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