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Probe pool for detecting ovarian cancer molecular subtypes, and preparation, application and use method thereof

A molecular typing, ovarian cancer technology, applied in biochemical equipment and methods, microbial determination/examination, DNA/RNA fragments, etc. High throughput effect

Active Publication Date: 2022-04-29
SUZHOU FANGKE BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Surgery and chemotherapy can cure most early stage patients, but cannot save the lives of many advanced stage patients

Method used

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  • Probe pool for detecting ovarian cancer molecular subtypes, and preparation, application and use method thereof

Examples

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Effect test

preparation example Construction

[0036] A method for preparing a probe pool for detecting molecular typing of ovarian cancer, comprising the following steps:

[0037] (1) Select multiple gene regions in claims 1, 2, and 3, and design probes according to the principle of base complementarity, and select hg19 for the human genome library;

[0038] (2) The region probe sequence in claims 1 and 2 needs to avoid the repeated region position;

[0039] (3) Probes need to remove probes with matching times greater than 1-200 on the human genome hg19;

[0040] (4) The size of the probe adjustment window is between 10-30nt;

[0041] (5) After the length of the probe is adjusted to 100-160 nt, the final probe sequence is formed, and then the probe is synthesized to form a probe pool.

[0042] The probe design principle of the present invention has a certain degree of advancement, and the influence between two different molecular types has been fully considered in the design, mainly due to the interference between diffe...

Embodiment 1

[0053] This embodiment provides a DNA probe pool, which contains 223 oligonucleotide probes, the nucleotide sequences of which are respectively shown in SEQ ID NO.1-SEQ ID NO.223.

[0054] Its preparation method comprises the following steps:

[0055] (1) Select multiple gene regions, and design probes according to the principle of base complementarity, and select hg19 for the human genome library;

[0056] (2) The probe sequence needs to avoid the position of the repeating region. The first choice should be to select a position that does not exceed 30-50 nt upstream and downstream of the repeating region for displacement. If the probe cannot be obtained effectively after repeated adjustments, increase the window size. more than 100nt;

[0057] (3) Probes need to remove probes that match more than 1-200 times on the human genome hg19, and mark them as risk probes to form a risk probe pool. In principle, the risk probe pool is not used for subsequent analysis and detection, bu...

Embodiment 2

[0062] This example verifies the detection accuracy of the DNA probe pool prepared in Example 1 for molecular typing of ovarian cancer. The specific methods and steps are as follows:

[0063] Use 100ng of DNA for each mutation standard;

[0064] KAPA Hyper DNA Library Preparation Kit was used for next-generation sequencing library construction, and the library construction operation was carried out in full accordance with the operation manual of the kit;

[0065] Take 200ng of a single library from each library and mix them into a total of 1μg mixed library;

[0066] Hybridize the mixed library with the probe pool provided by the present invention, the hybridization capture reagent uses IDT xGen DNACapture Kit, and the operation steps are completely carried out according to the operation manual of the kit;

[0067] The hybridized and captured library was sequenced on the NovaSeq6000 machine according to the standard procedure, and the total output of the mixed library was ab...

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Abstract

The invention discloses a probe pool for detecting ovarian cancer molecular subtypes as well as preparation, application and a use method thereof. The preparation method comprises the following steps: 1) designing a probe for I-type ovarian cancer; (2) designing a probe for the type II ovarian cancer; the design of the probe for I-type ovarian cancer typing comprises a combination of a plurality of regions of BRAF, KRAS, NRAS, ARAF, HRAS, PTEN, POLE and POLD genes; the design of the probe for type II ovarian cancer typing comprises combination of a plurality of regions of genes TP53, BRCA1, BRCA2, ATM, ATR, CCND1, CCNE1, ARID1A, CDK4, CDK6, CDK12 and PIK3CA, and the probe has the advantages that two common ovarian cancer molecular typing can be detected at one time by combining and using a targeted next-generation sequencing method, the flux is high, the precision is high, the cost is low, the timeliness is better, and the probe is suitable for clinical application. And a technical guarantee is provided for ovarian cancer molecular typing diagnosis, auxiliary medication guidance and prognosis prediction monitoring.

Description

technical field [0001] The invention relates to the field of molecular typing of ovarian cancer, in particular to a probe pool for detecting molecular typing of ovarian cancer and its preparation, application and usage methods. Background technique [0002] Ovarian malignant tumor is one of the common malignant tumors of female reproductive organs, and its incidence rate ranks third after cervical cancer and endometrial cancer. Among them, epithelial tumors account for 50%-70% of primary ovarian tumors, and their malignant types account for 85%-90% of ovarian malignant tumors. Surgery and chemotherapy can cure most early-stage patients, but fail to save the lives of many advanced-stage patients. Ovarian cancer is sensitive to cisplatin. Targeted therapy (such as anti-angiogenic drugs or poly ADP-ribose polymerase inhibitors) can also improve the survival rate of patients. Epithelial ovarian cancer affects 220,000 women worldwide each year. In the UK, 7,000 women are affe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12Q1/6869C12Q1/6806C12N15/11
CPCC12Q1/6886C12Q1/6869C12Q1/6806C12Q2600/112C12Q2600/118C12Q2600/106C12Q2535/122C12Q2525/113C12Q2563/131
Inventor 魏冬凯闻彬浦宇
Owner SUZHOU FANGKE BIOTECHNOLOGY CO LTD
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