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pik3ca gene mutation detection system and its kit

A gene and system technology, applied in the biological field, can solve problems such as poor efficacy, and achieve the effects of less primers and probes, high sensitivity and high throughput

Active Publication Date: 2019-07-05
苏州英泽生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Recent studies have also confirmed that PIK3CA gene mutations can cause breast cancer patients to be resistant to Her2-targeted drug injection Trastuzumab (Trastuzumab, trade name Herceptin / Herceptin), resulting in poor efficacy

Method used

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  • pik3ca gene mutation detection system and its kit

Examples

Experimental program
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Embodiment 1

[0047] 1. The composition of the kit.

[0048] The PIK3CA gene mutation detection kit of this embodiment includes: PCR reaction solution, universal forward primer, quality control reverse primer, detection reverse primer, probe (probe), amplification blocking primer (blocker), positive control, Negative control and blank control, as shown in Table 1.

[0049] Table 1 Kit composition table

[0050]

[0051]

[0052] The description of each component of the kit in the above Table 1 is as follows:

[0053] 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 Taq DNA polymerase at a concentration of 5U / μl, and the final concentration in the reaction system is 0.05U / μl. ...

Embodiment 2

[0117] 1. The composition of the kit.

[0118] The PIK3CA gene mutation detection kit of this embodiment includes: PCR reaction solution, universal forward primer, quality control reverse primer, detection reverse primer, probe (probe), amplification blocking primer (blocker), positive control, Negative control and blank control, as shown in Table 8.

[0119] Table 8 Kit composition table

[0120]

[0121]

[0122] The description of each component of the kit in the above Table 8 is as follows:

[0123] 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 Taq DNA polymerase at a concentration of 5U / μl, and the final concentration in the reaction system is 0.05U / μl. 10...

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Abstract

The invention discloses a PIK3CA gene mutation detection system and a kit thereof. The PIK3CA gene mutation detection system comprises primers, a probe and an amplification blocking primer, wherein the primers include a general forward primer, detecting reverse primers and a quality-control reverse primer. The detecting reverse primers include the detecting reverse primer for the E542K mutation site of a PIK3CA gene, the detecting reverse primer for the E545K mutation site of the PIK3CA gene, the detecting reverse primer for the H1047R mutation site of the PIK3CA gene and the detecting reverse primer for the H1047L mutation site of the PIK3CA gene. The 5' end of a probe sequence is provided with a fluorescent marker, and the 3' end of the nucleotide sequence of the probe is provided with another fluorescent marker. The 3' end of the nucleotide sequence of the amplification blocking primer is provided with a phosphorylated modifier. The PIK3CA gene mutation detection system and the kit of the system can detect the E542K, E545K, H1047R or H1047L mutation site of the PIK3CA gene and are high in sensitivity, strong in specificity and fast and convenient to use.

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] Recent ideas in cancer research suggest that cancer depends on driver genes for cancer maintenance, and that these driver genes are the best targets for therapy. The driver gene is an important gene related to the occurrence and development of cancer, and is the most important cause of cancer; when the driver gene is mutated, it will "drive" the cancer cells. At present, the analysis of driver genes in patients with non-small cell lung cancer according to histological type and smoking status shows that the driver genes of lung cancer include EGFR, KRAS, BRAF, PIK3CA and EML4-ALK, and the occurrence and progression of tumors are related to these genes; among them, more and more Many studies have found that driver gene mutations exist in lung squamous cell carcino...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6858C12N15/11
Inventor 赵新泰王明张长顺
Owner 苏州英泽生物医药科技有限公司
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