Real-time fluorescent quantitative PCR (polymerase chain reaction) method of TAQMAN probe with standard plasmids built by tree shrew IL-6 (interleukin 6)

Abstract

The invention discloses a real-time fluorescent quantitative PCR (polymerase chain reaction) method of a TAQMAN probe with standard plasmids built by tree shrew IL-6 (interleukin 6). The real-time fluorescent quantitative PCR method comprises the steps of cloning and sequencing a tree shrew IL-6 gene segment, by taking the cloned tree shrew 1L-6 gene segment as a target, designing a TAQMANPCR primer and a probe, designing the annealing temperature of the primer, the primer concentration and the probe concentration when the optimal condition is found out in an experiment; purifying the tree shrew IL-6 plasmids and computing the copy number of the IL-6 gene segment, diluting the calculated tree shrew IL-6 plasmids by 10-fold gradient, preparing positive plasmids of which the copy number is 10<5> to 10<8> copies to build a standard curve, and detecting the sensitivity.

Description

technical field [0001] The invention belongs to the field of molecular biology, in particular to a TAQMAN probe real-time fluorescent quantitative PCR detection method based on the cloned tree shrew IL-6 plasmid as a standard product. Background technique [0002] Interleukin 6 (Interleukin 6, IL-6), also known as β2-interferon, is a cytokine secreted by lymphoid and non-lymphoid cells with various biological functions, which can stimulate a variety of cell proliferation, Differentiation, and plays an important role in the body's immune response, bone marrow hematopoiesis, autoimmunity and body defense. In the anti-infection process, IL-6 is the main pro-inflammatory cytokine, which can promote the differentiation of CD4 T cells and promote T cells to produce IL-2 and express IL-2 receptor. In addition, IL-6 can also promote the differentiation of plasma cells and the production of antibodies by B cells. [0003] The tree shrew (Tupaia belangeri) is a small mammalian climb...

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Problems solved by technology

Some studies have also found that IL-6 plays a very important role in promoting the chronicity of HCV infection and liver cell damage; similar changes in the serum of patients with severe hepatitis B; also found that in severe hand, foot and mouth disease In the serum of children, IL-6 is significantly higher than that of the normal control group, which may be involved in the pathological process of severe hand, foot and mouth disease. Evaluation indicators of model immune cytokines

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