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IDH1/2 gene mutation detection system and kit thereof

A technology of IDH1 and gene, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of increased risk of malignant brain tumors, and achieve the effect of less primers and probes, high sensitivity, and high throughput

Active Publication Date: 2016-03-02
苏州元德友勤医学检验所有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Excessive accumulation of 2HG has been shown to lead to an increased risk of malignant brain tumors in patients with congenital abnormalities in 2HG metabolism

Method used

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  • IDH1/2 gene mutation detection system and kit thereof
  • IDH1/2 gene mutation detection system and kit thereof
  • IDH1/2 gene mutation detection system and kit thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] 1. The composition of the kit.

[0051] The IDH1 / 2 gene mutation detection kit of this embodiment includes: PCR reaction solution, quality control forward primer, detection forward primer, universal reverse primer, probe (probe), amplification blocking primer (blocker), positive The control, negative control and blank control are shown in Table 1.

[0052] Table 1 Kit composition table

[0053]

[0054]

[0055] The description of each component of the kit in the above Table 1 is as follows:

[0056] The PCR reaction solution is prepared from 10×PCR buffer, dNTPs and hot-start enzyme. 10×PCR buffer includes 100mM Tris-HCl, 500mM KCl and 15mM MgCl 2 , the pH value of the Tris-HCl buffer used to configure the PCR buffer is 8.3. dNTPs include dATP, dGTP, dCTP and dTTP, and the final concentration in the reaction system is 0.2mM. The hot start enzyme uses TaqDNA polymerase at a concentration of 5U / μl, and the final concentration in the reaction system is 0.05U / μl...

Embodiment 2

[0126] 1. The composition of the kit.

[0127] The IDH1 / 2 gene mutation detection kit of this embodiment includes: PCR reaction solution, quality control forward primer, detection forward primer, universal reverse primer, probe (probe), amplification blocking primer (blocker), positive Control, negative control and blank control, as shown in Table 8.

[0128] Table 8 Kit composition table

[0129]

[0130]

[0131] The description of each component of the kit in the above Table 8 is as follows:

[0132] The PCR reaction solution is prepared from 10×PCR buffer, dNTPs and hot-start enzyme. 10×PCR buffer includes 100mM Tris-HCl, 500mM KCl and 15mM MgCl 2 , the pH value of the Tris-HCl buffer used to configure the PCR buffer is 8.3. dNTPs include dATP, dGTP, dCTP and dTTP, and the final concentration in the reaction system is 0.2mM. The hot start enzyme uses TaqDNA polymerase at a concentration of 5U / μl, and the final concentration in the reaction system is 0.05U / μl. 1...

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Abstract

The invention discloses an IDH1 / 2 gene mutation detection system and a kit thereof. The IDH1 / 2 gene mutation detection system comprises primers, probes and an amplification blocking primer; the primers comprise the quality-control forward primers, the detection forward primers and the general reverse primers; the detection forward primers comprise the detection forward primer for a C394T mutation site of an IDH1 gene, the detection forward primer for a G395A mutation site of the IDH1 gene, the detection forward primer for a G419A mutation site of an IDH2 gene and the detection forward primer for a G515A mutation site of the IDH2 gene; the 5'end of a probe sequence is provided with a fluorescent mark, and the 3'end of the nucleotide sequence of the probe is provided with another fluorescent mark; the 3'end of a nucleotide sequence of the amplification blocking primer is provided with phosphorylation modification. According to the IDH1 / 2 gene mutation detection system and the kit thereof, the C394T and G395A mutation sites of the IDH1 gene and the G419A and G515A mutation sites of the IDH2 gene can be detected, the sensitivity and the specificity are high, and rapidness and convenience are achieved.

Description

technical field [0001] The invention relates to a gene mutation detection product and primers, probes and detection systems used in the product, belonging to the field of biotechnology. Background technique [0002] Isocitrate dehydrogenase (IDH) is an enzyme family that plays an important role in the tricarboxylic acid cycle. It not only plays an important role in energy metabolism, amino acid and vitamin synthesis, but also regulates the activity of this enzyme will directly Affect IDH or IDH substrates to participate in different biological pathways and exert different biological functions. In mammalian cells, IDH isozymes have three forms: NAD-dependent mitochondrial IDH, NADP-dependent mitochondrial IDH, and NADP-dependent cytosolic IDH. [0003] The gene encoding NADP-dependent human isocitrated dehydrogenase 1 (isocitratedehydrogenase1, IDH1) is located on chromosome 2q33.3, and its transcribed mRNA is 2339 bp long, contains 10 exons, and encodes isocitrate dehydroge...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 赵新泰王明张长顺
Owner 苏州元德友勤医学检验所有限公司
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