Method for detecting CLC gene expression quantity in nasal cavity exfoliated cells and application of method

A technology of gene expression and exfoliated cells, which is applied to the detection of CLC gene expression in nasal exfoliated cells, can solve the problems of non-healing mucosal pathological biopsy, failure to reflect the overall picture of the tissue, and increase the risk of infection in patients

Pending Publication Date: 2018-12-11
张罗 +4
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The disadvantages of pathological biopsy of nasal mucosa are as follows: 1. It is an invasive examination: it increases the risk of infection for patients, and is not suitable for people with low immunity such as children and the elderly; nasal cavity bleeding often causes fear and worry in patients
2. It is difficult to obtain real-time dynamic change information of the disease: pathological biopsy of the healing mucosa cannot be performed after the operation
However, clinical data show that the pathological classification of chronic sinusitis with nasal polyps will vary with the outcome of drug treatment, surgical treatment, etc. Pathological biopsy results before treatment cannot represent the characteristics of all disease stages
3. Time-consuming and increase the cost of medical treatment, usually 3-4 working days from obtaining tissue samples to obtaining pathological results
Due to the inability to obtain the results on the same day or the next day, extra transportation, accommodation, and registration fees will be incurred for out-of-town patients seeking medical treatment, which increases the cost of medical treatment
4. There are certain human errors, and the number of inflammatory cells counted by different pathologists may be different, which affects the judgment of polyp typing
5. Histopathological sections are relatively limited, and can only reflect the inflammatory state of the specimen at the section position, but cannot reflect the whole picture of the tissue, which may cause misdiagnosis
6. Each film needs to be manually counted by pathologists, which is difficult to operate in batches
However, the prior art has not yet disclosed an effective method for detecting CLC gene expression in nasal exfoliated cells, so as to better obtain the CLC gene content in nasal exfoliated cells

Method used

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  • Method for detecting CLC gene expression quantity in nasal cavity exfoliated cells and application of method
  • Method for detecting CLC gene expression quantity in nasal cavity exfoliated cells and application of method
  • Method for detecting CLC gene expression quantity in nasal cavity exfoliated cells and application of method

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preparation example Construction

[0059] The preparation method of the DNase reaction solution comprises the following steps: taking DNase buffer solution, recombinant DNase, and double distilled water without RNase and mixing to obtain the DNase reaction solution. Preferably, the preparation method of the DNase reaction solution comprises the following steps: take 5 μL of 10×DNase buffer solution, 4 μL of recombinant DNase, and 41 μL of RNase-removed double-distilled water and mix to obtain the DNase reaction solution.

[0060] As a preference, when the genome content is low or the initial amount of material is low when the RNA is extracted, the method for extracting RNA from nasal exfoliated cells further includes the following steps: in the step 1, the nasal exfoliated cells are lysed After being added to the cell lysate, it is first added to the genomic DNA adsorption column to obtain the filtrate, and then an equal volume of ethanol is added to the filtrate.

[0061] Preferably, in order to obtain a highe...

Embodiment 1

[0091] Sample collection and processing:

[0092] After rinsing the nasal cavity with normal saline, a patient pressed a brush (manufactured by Copan) on the surface of the nasal polyp for 30 seconds under the nasal endoscope, rotated it 3 to 4 times, and brushed the surface of the polyp. ℃ for short-term storage (not exceeding 24 hours), or transfer to below -20 ℃ for long-term storage.

[0093] A method for detecting CLC gene expression in nasal cavity exfoliated cells, comprising the following steps:

[0094] Step 1: Extract RNA from nasal exfoliated cells:

[0095] Step 1: Take the genomic DNA adsorption column and place it in a 2mL collection tube, dissolve the nasal exfoliated cells in 300 μL of cell lysate and add them to the genomic DNA adsorption column, take the filtrate, and add an equal volume of 70% Ethanol, mixed evenly, was added to the RNA purification column, 12000 rpm, centrifuged for 1min, the filtrate was removed, and the RNA purification column was place...

Embodiment 2

[0106] Sample collection and processing:

[0107] After rinsing the nasal cavity with normal saline, a patient pressed a brush (manufactured by Copan) on the surface of the nasal polyp for 30 seconds under the nasal endoscope, rotated it 3 to 4 times, and brushed the surface of the polyp. ℃ for short-term storage (not exceeding 24 hours), or transfer to below -20 ℃ for long-term storage.

[0108] A method for detecting CLC gene expression in nasal cavity exfoliated cells, comprising the following steps:

[0109] Step 1: Extract RNA from nasal exfoliated cells:

[0110] Step 1: Take the genomic DNA adsorption column and place it in a 2mL collection tube, dissolve the nasal exfoliated cells in 100 μL of cell lysate, add them to the genomic DNA adsorption column, centrifuge at 12,000 rpm for 60 seconds, and take the filtrate. Add an equal volume of 70% ethanol to the filter, mix well and add to the RNA purification column, centrifuge at 12000 rpm for 1 min, remove the filtrate,...

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Abstract

The invention provides a non-diagnostic method for detecting a CLC gene expression quantity in nasal cavity exfoliated cells and application of the method. The method comprises the following steps ofextracting RNA from the nasal cavity exfoliated cells, performing reverse transcription on total RNA to obtain cDNA, adopting quantitative polymerase chain reaction to perform real-time fluorescent quantitative PCR (polymerase chain reaction) amplification on a CLC gene and a reference gene in the cDNA by adopting a specific primer of the CLC gene and a specific primer of the reference gene respectively, and calculating the CLC gene expression quantity based on a detection result of an amplification product. For the method provided by the invention, by taking the effectively screened CLC geneas a biomarker, a detection method of the gene expression quantity of the CLC gene is provided, so that calculation of the CLC gene expression quantity in the nasal cavity exfoliated cells is realized, and the CLC gene expression quantity can be effectively acquired; and in addition, the provided method is simple and rapid, is good in sensibility and good in repeatability and is suitable for beingwidely popularized and applied.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a method for detecting the expression of CLC gene in exfoliated cells of the nasal cavity. Background technique [0002] Chronic rhinosinusitis with nasal polyps (CRSwNP) is a chronic inflammation of the sinus mucosa, and physical examination shows the formation of polyps in the nasal cavity or middle nasal passage. Common symptoms of CRSwNP are nasal congestion, runny nose or backflow, hyposmia, facial fullness or pressure, and last for more than 12 weeks. The prevalence rate is about 0.5-4%. CRSwNP is often accompanied by asthma and allergic rhinitis. It has been reported that 7% of asthmatic patients suffer from CRSwNP, while 26-48% of CRSwNP suffer from asthma. The pathogenesis of CRSwNP is still uncertain. The destruction of mucosal epithelial cells, the imbalance of host immune system and the invasion of pathogenic microorganisms may be the main causes of i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6851C12Q1/6883
CPCC12Q1/6851C12Q1/6883C12Q2600/166C12Q2561/113C12Q2563/107C12Q2531/113
Inventor 张罗王成硕闫冰刘畅
Owner 张罗
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