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ASPL-TFE3 fused renal carcinoma gene probe as well as application of kit thereof

A technology of gene probe and kit, which is applied in the application field of ASPL-TFE3 fusion kidney cancer gene probe and its kit, can solve the problems of limiting RT-PCR, inability to accurately diagnose kidney cancer, and not being suitable for wide application, etc. To achieve the effect of rapid diagnosis, high success rate and simple diagnosis

Inactive Publication Date: 2015-03-04
THE AFFILIATED DRUM TOWER HOSPITAL MEDICAL SCHOOL OF NANJING UNIV
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  • Abstract
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Problems solved by technology

For karyotype analysis, tumor cells must be cultured to the mitotic stage, and then G-banding and other techniques are used to analyze the karyotype of tumor cells. This method is cumbersome and time-consuming and is not suitable for clinical application.
Similarly, considering the rarity of ASPL-TFE3 fusion RCC and the complexity of the PCR test process, it is impossible to routinely use RT-PCR to detect ASPL-TFE3 fusion genes in all RCC tissues, and when ASPL-TFE3 fusion is suspected In advanced renal cancer, RNA can only be extracted from paraffin sections. The rapid degradation of RNA and many other factors limit the application of RT-PCR, and the technology is complicated and not suitable for wide application.
These limitations lead to the inaccurate and timely diagnosis of ASPL-TFE3 fusion RCC, which affects the judgment of clinicians on the prognosis and postoperative management of patients with ASPL-TFE3 fusion RCC

Method used

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  • ASPL-TFE3 fused renal carcinoma gene probe as well as application of kit thereof
  • ASPL-TFE3 fused renal carcinoma gene probe as well as application of kit thereof
  • ASPL-TFE3 fused renal carcinoma gene probe as well as application of kit thereof

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Experimental program
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Effect test

Embodiment 1

[0041] Step 1: Preparation of Gene Probes:

[0042] Find the bacterial artificial chromosome (BAC clone) corresponding to the X chromosome TFE3 gene and ASPL gene through http: / / genome.ucsc.edu / , select the corresponding BAC clone fragments respectively, and control the complete coverage of the BAC clone fragments selected on the same side relative to the gene , and the ipsilateral fragments have a certain sequence overlap with each other or the distance is less than 10Kb. According to the above requirements, the BAC clone fragments on the ASPL gene were selected as RP11-634L10 (chr17: 79796813-79969288, the fragment length is about 172Kb), RP11-51H16 (chr17: 79931578-80097452, the fragment length is about 166Kb) and RP11-475F12 ( chr17: 80032268-80216871, the fragment length is about 185Kb); the BAC clone fragment on the TFE3 gene is CTD-2311N12 (chrX: 48713289-48923401, the fragment length is about 210Kb), CTD-2516D6 (chrX: 48265836-48484403, The fragment length is about 21...

Embodiment 2

[0054] Example 2: ASPL-TFE3 Fusion Kidney Cancer Diagnostic Kit

[0055] Gene probe kit for ASPL-TFE3 fusion kidney cancer, the kit is composed of probe hybridization solution and 4', 6-diamidino-2-phenylindole counterstaining agent, characterized in that:

[0056] (1) The combination of RP11-634L10, RP11-51H16, and RP11-475F12 located on the ASPL gene on chromosome 17, marked with a red fluorescent signal; Combination of CTD-2516D6, CTD-2312C1, CTD-2248C21 and RP11-959H17, labeled with green fluorescent signal;

[0057] (2) The probe hybridization solution is prepared by mixing the probe with Human Cot-1 DNA, hybridization buffer, and purified water in proportion, and it needs to be stored in a dark place at -20°C;

[0058] (3) 4′,6-diamidino-2-phenylindole counterstain is mainly used for nuclear staining.

[0059] The combination of RP11-634L10, RP11-51H16 and RP11-475F12 located on the ASPL gene of chromosome 17 is marked with a red fluorescent signal; located on the TFE3...

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Abstract

The invention relates to an ASPL-TFE3 fused renal carcinoma gene probe as well as an application of a kit thereof. Cloned fragments selected by the gene probe are respectively a combination of RP11-634L10, RP11-51H16 and RP11-475F12 and a combination of CTD-2311N12, RP11-416B14, CTD-2522M13, CTD-2516D6, CTD-2312C1, CTD-2248C21 and RP11-959H17. By using the ASPL-TFE3 fused renal carcinoma gene probe, the defects of complexity, time consumption and clinic application unsuitability of RT-PCR and cell karyotyping methods applied in the past are overcome. According to the ASPL-TFE3 fused renal carcinoma gene probe, a unique ASPL-TFE3 fused gene in ASPL-TFE3 fused renal carcinoma is detected by using an FISH technology, so that the renal carcinoma is diagnosed; the gene probe is high in accuracy rate, specificity and success rate, strong in fluorescence signal, simple and rapid in operation and rapid in diagnosis when applied and can be applied to paraffin section; and due to the application of the ASPL-TFE3 fused renal carcinoma gene probe, the specimen detecting range is widened, a novel method for accurately, reliably, simply and conveniently diagnosing the ASPL-TFE3 fused renal carcinoma is established, and the precedent that the ASPL-TFE3 fused renal carcinoma is detected by virtue of FISH is created.

Description

technical field [0001] The invention belongs to the application field of fluorescent in situ hybridization probes, in particular to the application of ASPL-TFE3 fusion kidney cancer gene probes and kits thereof. Background technique [0002] Xp11.2 translocation / TFE3 gene fusion-related renal cell carcinoma is more common in adolescents, accounting for about 1 / 3 of renal cancer in children, and 15% of renal cancer under the age of 45. The prognosis is poor, and the prognosis of adult patients is worse than that of children. It is mainly related to pathological type, tumor grade and stage, whether there is distant metastasis, etc. Xp11.2 translocation / TFE3 gene fusion correlation is not effective for conventional immunotherapy, and it is not sensitive to radiotherapy and chemotherapy. Surgery is the main treatment method, and patients are prone to lymph node and distant metastasis. Only targeted therapy has been proved to have a certain effect. effect. [0003] ASPL-TFE3 fu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12Q1/68
Inventor 甘卫东陈显成何健杨军郭宏骞贾瑞鹏
Owner THE AFFILIATED DRUM TOWER HOSPITAL MEDICAL SCHOOL OF NANJING UNIV
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