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Primer and probe combinations, kits and applications for early screening of bladder cancer

A primer probe and bladder cancer technology, which is applied in the field of primer and probe combinations for early screening of bladder cancer, can solve the problems of low sensitivity and poor specificity of bladder cancer detection, and achieve simple and easy detection, objective interpretation, and improved Check the effect of sensitivity

Active Publication Date: 2021-11-30
北京求臻医疗器械有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In view of the deficiency of the above-mentioned background technology, the present invention provides a combination of primers and probes, kits and applications for early screening of bladder cancer, which solves the problem of low sensitivity and poor specificity of bladder cancer detection, and avoids problems caused by cystoscopy. The problem of complications

Method used

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  • Primer and probe combinations, kits and applications for early screening of bladder cancer
  • Primer and probe combinations, kits and applications for early screening of bladder cancer
  • Primer and probe combinations, kits and applications for early screening of bladder cancer

Examples

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Effect test

Embodiment 1

[0045] Example 1 Kit for Early Screening of Bladder Cancer

[0046] Including Qiagen urine free DNA extraction kit, HinP1I, HpaII, AciI, HpyCH4IV methylation-sensitive endonuclease system, 4 pairs of target-specific primers and internal reference primers, 4 pairs of specific primer probes, internal reference primer probes and internal reference primers probe;

[0047] Wherein the sequences of the 4 pairs of target-specific primers are as follows:

[0048] Seq ID NO.1: F-CCGCGCTCACGTCGGTTCC,

[0049] Seq ID NO.2: R-CCCCCTACTTCCCCAGCTCACGA;

[0050] Seq ID NO.3: F-TCCTATCCTCATCGGTTCTCGTACT,

[0051] Seq ID NO.4: R-AATTGTCCAACGAATGCACCTG;

[0052] Seq ID NO.5: F-CCCGCACACTACTCCAAATCAA,

[0053] Seq ID NO.6: R-GCAAAATCGAGGGCAGCTGAAG;

[0054] Seq ID NO.7: F-CAGCGCGTGTCCTTCTTCGCCTG,

[0055] Seq ID NO.8: R-AGTTCTCGAGCGTGCCACCGTTG;

[0056] The sequences of internal reference primers are as follows:

[0057] Seq ID NO.9: F-GACCCGCGCTTCGACACCG,

[0058] Seq ID NO.10: R-CCAG...

Embodiment 2

[0066] Example 2 Method for detecting changes in ctDNA methylation level

[0067] 1. Take 10 mL of urine and use Qiagen Urine Free DNA Extraction Kit (Cat: 954154) to extract free cfDNA;

[0068] 2. Take 20 ng cfDNA and 20 μL HinP1I, HpaII, AciI, HpyCH4IV four methylation-sensitive endonuclease systems (final concentration 10 U / μL), incubate at 37°C for 16 h, and inactivate the enzyme at 80°C for 20 min;

[0069] 3. Use all the incubated products as templates, 4 pairs of target-specific primers (50 nM each) plus 1 internal reference primer (10 nM) to configure the system, set the following table 1 to set the program and perform the amplification program in a common PCR instrument ;

[0070] Table 1

[0071]

[0072] 4. Take 1 μL of the amplification product from the previous step as a template, add 4 pairs of target-specific primers (0.25 μM each), 4 specific primer probes (0.1 μM each), plus 1 internal reference primer (0.05 μM) and internal reference Primer probe (0.02...

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Abstract

The present invention relates to the field of molecular biology technology, in particular to a combination of primers and probes, kits and applications for early screening of bladder cancer, and a combination of primers and probes for early screening of bladder cancer, including the combination of primers and probes for early screening of bladder cancer. 4 pairs of target-specific primers and corresponding internal reference primers and specific primer probes related to ylation sites. Compared with the prior art, the detection based on DNA methylation in the present invention is simple, easy to perform, objective in interpretation, avoids the subjectivity of human interpretation results, and improves the accuracy rate; at the same time, the detection method of the present invention is non-invasive and can Avoid complications caused by cystoscopy, improve patient compliance, and be used for early screening, early diagnosis, curative effect evaluation and tracking, and prognosis evaluation of tumors.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to a primer and probe combination, kit and application for early screening of bladder cancer. Background technique [0002] Bladder cancer is one of the most common malignant tumors of the urinary system. In 2014, the incidence of bladder cancer in China was 78,000 people per year, and the incidence rate is increasing year by year. Bladder cancer has the characteristics of high incidence and easy recurrence. Hematuria is a common clinical symptom of bladder cancer, and bladder cancer is detected in about 17% of patients with hematuria. Currently, the main diagnostic methods for bladder cancer include shadow cystoscopy, urine exfoliation cytology, urine FISH and tumor markers. Among them, cystoscopy combined with biopsy histopathology is the gold standard for cystoscopic diagnosis, but this examination method is invasive, prone to complications, and patient compliance is ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6886C12Q1/6851C12N15/11
CPCC12Q1/6886C12Q1/6851C12Q2600/154C12Q2600/166C12Q2521/331C12Q2531/113C12Q2545/101
Inventor 张怡然王冰商宇红牛春影顾丽清王天阳
Owner 北京求臻医疗器械有限公司
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