Primer pairs, probe, kit and one-step detecting method for detecting tumor fusion gene EWS-FLI1

A technology that combines genes and detection methods, applied in the field of genetic engineering, can solve the problems of expensive probes, cumbersome, time-consuming detection, etc., and achieve the effect of objective and accurate detection results and simple operation methods

Inactive Publication Date: 2018-11-13
青岛普泽麦迪生物技术有限公司
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Problems solved by technology

However, FISH detection takes a long time, the cost of the probe is relatively expensive, and professionally trained technicians are required to perform cumbersome operations, the result interpretation is time-consuming and there are subjective errors, etc.

Method used

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  • Primer pairs, probe, kit and one-step detecting method for detecting tumor fusion gene EWS-FLI1
  • Primer pairs, probe, kit and one-step detecting method for detecting tumor fusion gene EWS-FLI1
  • Primer pairs, probe, kit and one-step detecting method for detecting tumor fusion gene EWS-FLI1

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Embodiment 1

[0030] Material Description:

[0031] Template: The sample provided in this example is Total RNA extracted from tumor tissue, but because the incidence of Ewing sarcoma is very low, it is extremely difficult to obtain samples, therefore, the Ewing sarcoma cell line A673 ( Total RNA of CRL-1598) was used as a positive control for type I fusion, and Total RNA from tumor tissue of Ewing sarcoma patient Z63 was used as a positive control for type II fusion.

[0032] 1. Sample one-step RT-qPCR experiment

[0033] PCR experiments using NEB Universal Probe One-Step RT-qPCR Kit, the specific steps include:

[0034] 1) will Universal ProbeOne-Step Reaction Mix (2x) is thawed at room temperature, gently mixed upside down and centrifuged for a short period of time, for use, and always keep away from light during use;

[0035] 2) Prepare the PCR reaction solution of the sample

[0036] Prepare sample EWS-FLII type fusion PCR reaction solution and EWS-FLI1 type II fusion PCR reacti...

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Abstract

The invention provides primer pairs, probe, kit and one-step detecting method for detecting tumor fusion gene EWS-FLI1, belongs to the technical field of gene engineering, and can solve the problems that existing fusion gene EWS-FLI1 detection is complex in operation and time-consuming in result interpretation and has subjective errors. The primer pairs comprise a forward primer EWS_F as shown inSEQ No. 1 and a reverse primer FLI1_Exon 7&8 R as shown in SEQ No. 2 which are used for fusion gene EWS-FLI1 I fusion, and a forward primer EWS_F as shown in SEQ No. 1 and a reverse primer FLI1_Exon 6&7 R as shown in SEQ No. 4 which are used for fusion gene EWS-FLI1 II fusion. The primer pairs are applicable to the quantitative detection of the fusion gene EWS-FLI1 and thus can be effectively usedin the detection of Ewing sarcoma family tumor.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and particularly relates to a primer pair, a probe, a kit and a one-step detection method for tumor fusion gene EWS / FLI1 detection. Background technique [0002] Ewing sarcoma family tumors (ESFT) are a group of poorly differentiated and highly malignant small round cell tumors, including Ewing sarcoma (Ewingsarcoma, ES), Askin tumor and primitive neuroectodermal tumor (PNET). The median age of the patients was 15 years, and more than half of the patients were adolescents. The annual incidence of ESFT in children under the age of 15 is 3 / million people, and about 30% of the patients are found to have metastases in other parts such as lung and bone at the time of diagnosis. [0003] At present, seven chromosomal translocations have been found in ESFT patients, and the t(11;22)(q24;q12) translocation, that is, the fusion of the FLI1 gene on chromosome 11 q24 and the EWS gene on chromos...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886
Inventor 姚斐宫小明马荣桧金超
Owner 青岛普泽麦迪生物技术有限公司
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