Preparation method for probes related to breast cancer molecular markers and application of same

A probe and gene probe technology, which is applied to the preparation of breast cancer molecular marker-related probes and their application fields, can solve the problems of decreased curative effect, poor patient prognosis, shortened recurrence-free survival period, etc., and achieves simple operation and improved Detection rate and effect of accurate breast cancer typing

Active Publication Date: 2012-04-04
DAAN GENE CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

TOP2A gene is involved in the replication of breast cancer cells. Patients with TOP2A gene abnormality have a poor prognosis and shorter recurrence-free survival, especially those with TOP2A gene deletion have a worse prognosis
Gene amplification indicates the possibility of tumor recurrence or long-term curative effect decline

Method used

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  • Preparation method for probes related to breast cancer molecular markers and application of same
  • Preparation method for probes related to breast cancer molecular markers and application of same
  • Preparation method for probes related to breast cancer molecular markers and application of same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Embodiment 1: Preparation of HER2 gene probe

[0047] (1) Primer design and clone screening: HER2 gene is located in the 17q12 segment of human chromosome. NCBI Mapview database searches all clones containing HER2 gene, and screens out the clone containing this gene, numbered RP11-909L6.

[0048] (2) Cloning culture and identification: Purchase the clone RP11-909L6, take 10 μl of the cloning bacteria solution and add it to 5 ml of TB culture solution (chloramphenicol resistance), and shake the bacteria in a shaker at 37°C for 8 to 12 hours; Add all the solution to 500ml TB culture solution (chloramphenicol resistance), shake the bacteria in a shaker at 37°C for 8-12 hours; the bacteria solution uses upstream primers

[0049] 5'-AGGGGAGAATAAATAAAATCTGTGG-3' and downstream primers

[0050] 5'-CAGGAGTGAGACACTCTCCATG-3' was amplified by PCR, and the amplification conditions were: 95°C for 5 minutes; (94°C for 30 seconds, 56°C for 30 seconds, 72°C for 45 seconds) × 40 cycle...

Embodiment 2

[0070] Embodiment 2: the preparation of TOP2A gene probe

[0071] (1) Primer design and clone screening: The TOP2A gene is located on the 17q21 segment of human chromosome, and all clones containing the TOP2A gene were searched in the NCBI Mapview database, and the clone containing the gene was screened out, numbered RP11-1029L16.

[0072] (2) Cloning culture and identification: purchase the clone RP11-1029L16, take 10 μl of the cloning bacteria solution and add it to 5ml TB culture solution (chloramphenicol resistance), and shake the bacteria in a shaking table at 37°C for 8-12 hours; Add all the solution to 500ml TB culture solution (chloramphenicol resistance), shake the bacteria in a shaker at 37°C for 8-12 hours; use the upstream primer 5'-ATTCAAAGCTGGATCCCTTT-3' and the downstream primer 5'-AGCTGTGACAAATGCCTGTA for the bacterial solution -3' was subjected to PCR amplification, and the amplification conditions were: 95° C. for 5 minutes; (94° C. for 30 seconds, 56° C. for...

Embodiment 3

[0092] Embodiment 3: the preparation of AGTR1 gene probe

[0093] (1) Primer design and clone screening: AGTR1 gene is located in the 3q24 segment of human chromosome. NCBI Mapview database searched all clones containing AGTR1 gene, and screened out the clone containing this gene, numbered RP11-366P9.

[0094] (2) Cloning culture and identification: purchase the clone RP11-366P9, take 10 μl of the cloning bacteria solution and add it to 5ml TB culture solution (chloramphenicol resistance), and shake the bacteria in a shaker at 37°C for 8 to 12 hours; Add all the solution to 500ml TB culture solution (chloramphenicol resistance), shake the bacteria in a shaker at 37°C for 8-12 hours; use the upstream primer 5'-TTGCATTATTTTCAAAGCCCTT-3' and the downstream primer 5'-TCCTACCAGCTTAGGCCAGA for the bacterial solution -3' was subjected to PCR amplification, and the amplification conditions were: 95° C. for 5 minutes; (94° C. for 30 seconds, 56° C. for 30 seconds, 72° C. for 45 seconds...

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Abstract

The invention relates to a preparation method for probes related to breast cancer molecular markers and to preparation of a breast cancer fluorescence in situ hybridization detection kit by using the probes. The breast cancer fluorescence in situ hybridization detection kit can be prepared from HER2, TOP2A and AGTR1 gene probes prepared by using the method provided in the invention, human chromosome 17 counting probe, hybridization buffer, unlabelled competitive DNA and DAPI counter strain; application of the kit enables the detection rate of breast cancers to be substantially improved and type sorting of breast cancers to be more accurate and provides guidance to formulation of an individual therapeutic schedule and selection of proper therapeutic drugs, thereby lowering down mortality, reducing recurrence risk and achieving the goal of optimizing the effect of diagnosis and treatment.

Description

technical field [0001] The present invention relates to a method for preparing breast cancer molecular marker-related probes, and also relates to using these probes to prepare breast cancer fluorescence in situ hybridization detection kits. The breast cancer molecular marker-related probes and corresponding kits of the present invention can be used It can be used to determine the type of breast cancer, especially to guide the formulation of an individualized treatment plan for breast cancer, evaluate the effect of treatment, monitor recurrence and metastasis, and estimate prognosis. Background technique [0002] Breast cancer is one of the most common malignant tumors in women, accounting for 7% to 10% of all kinds of malignant tumors in the whole body, second only to cervical cancer. Epidemiological survey found that 5% to 10% of breast cancer is familial, with obvious family genetic tendency. The incidence rate is higher in women between the ages of 40 and 60 and before a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10C12Q1/68G01N21/64
Inventor 李明何瑰陈华云
Owner DAAN GENE CO LTD
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