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Primer and probe set for diagnosis, detection or screening of digestive tract cancer

A digestive tract cancer and probe group technology, applied in the field of biomedicine, can solve the problems of low patient acceptance, low early detection rate of digestive tract cancer, and difficulty in popularization

Active Publication Date: 2018-11-13
SUZHOU VERSABIO TECH INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The current early screening method for gastrointestinal cancer is mainly endoscopy, but because this method requires professional operators to operate, it is difficult to popularize, and as an invasive inspection method, the acceptance of patients is low, which has led to The early detection rate of digestive tract cancer in China is much lower than that of developed countries

Method used

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  • Primer and probe set for diagnosis, detection or screening of digestive tract cancer
  • Primer and probe set for diagnosis, detection or screening of digestive tract cancer
  • Primer and probe set for diagnosis, detection or screening of digestive tract cancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] The genome sequence of a cell line known to be fully methylated with the SFRP2 gene was used as a DNA template. The DNA template was treated with bisulfite before the PCR reaction, and the ammonium bisulfite solution with a bisulfite concentration of 8M was used. The reaction conditions were: Heat at 70°C for 60 minutes, then use the Axygen gel purification kit to purify to obtain the converted DNA template, and then perform PCR.

[0027] Using SEQ ID NO.1, SEQ ID NO.17 and SEQ ID NO.37, carry out fluorescent quantitative PCR reaction, wherein the fluorescence labeled by SEQ ID NO.37 is ROX; the PCR system and reaction conditions are shown in Table 2 and Table 3.

[0028] The PCR component of table 2 embodiment 1

[0029] components

Final concentration

PCR reaction buffer

1X

DNA polymerase

0.1U / μL

dNTPs mixture

0.2mM

Mg2+

5mM

SEQ ID NO:1

0.2μM

SEQ ID NO: 17

0.2μM

SEQ ID NO: 37

0.1μM...

Embodiment 2

[0035] Colorectal cancer tissue and matched paracancerous tissue were used as detection objects, DNA was extracted with Qiagen’s DNeasy Blood&Tissue Kit, and then treated with 10M bisulfite, the bisulfite solution used was 6M ammonium bisulfite, A mixed solution of 3M sodium bisulfite and a mixed solution of 1M anhydrous sodium sulfate were heated at 80° C. for 40 minutes under the reaction conditions, and PCR was performed after purification using the Axygen gel purification kit.

[0036] Using SEQ ID NO.2, SEQ ID NO.22 and SEQ ID NO.31, wherein the fluorescence labeled by SEQ ID NO.31 is JOE, a fluorescent quantitative PCR reaction was used to detect 16 cases of colorectal cancer tissue and its paracancerous tissue samples. The PCR system and reaction conditions are shown in Table 4 and Table 5.

[0037] The PCR component of table 4 embodiment 2

[0038] components

Final concentration

PCR reaction buffer

1X

DNA polymerase

0.08U / μL

dN...

Embodiment 3

[0043] The test samples were 10 cases of esophageal cancer serum cfDNA, 31 cases of gastric cancer serum cfDNA and 36 cases of serum cfDNA of patients without obvious gastrointestinal diseases, and the serum volume was 1 mL. After the cfDNA extraction kit of Suzhou Weishan Biotechnology Co., Ltd. was used for DNA extraction, 9M bisulfite was used for treatment. The bisulfite solution used was 9M ammonium bisulfite solution. minutes, followed by PCR after purification using the Axygen Gel Purification Kit.

[0044] Using SEQ ID NO.5, SEQ ID NO.23 and SEQ ID NO.34, wherein the fluorescence labeled by SEQ ID NO.34 is FAM, for fluorescent quantitative PCR reaction detection, the PCR reaction system and reaction conditions are shown in Table 6 and Table 7 As shown, the test results are shown in Table 8.

[0045] The PCR component of table 6 embodiment 3

[0046] components

Final concentration

PCR reaction buffer

1X

DNA polymerase

0.15U / μL

...

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Abstract

The invention belongs to the field of biomedicine and relates to a primer and probe set for diagnosis, detection or screening of digestive tract cancer. Primers include a forward primer and a reverseprimer, and probes are SERP2 probes, wherein the forward primer of SERP2 includes any one sequence of SEQ ID: 1-16, and the reverse primer of SERP2 includes any one sequence of SEQ ID: 17-30. Comparedwith the prior art, the primer and probe set has the beneficial effects of by extracting DNA in a biological sample, whether the biological sample contains methylated SFRP2 gene is detected by a fluorescent quantitative PCR technique, thereby determining whether or not a risk of digestive tract cancer exists; the SFRP2 methylation sequence can be detected very well, and the detection sensitivitycan reach 10 copies / reaction; the methylated SFRP2 gene can be used for clearly distinguishing cancer samples and normal samples of colorectal cancer, gastric cancer, esophageal cancer and other digestive tract cancer.

Description

technical field [0001] The invention belongs to the field of biomedicine, and in particular relates to a primer and probe set for diagnosis, detection or screening of digestive tract cancer, which is used for early diagnosis / detection of digestive tract tumors such as colorectal cancer, esophageal cancer and gastric cancer. Background technique [0002] Digestive tract tumors (esophageal cancer, gastric cancer, and colorectal cancer) are the most common malignant tumors. In 2016, the 2015 Chinese cancer statistics published in the journal "CA: A Cancer Journal for Clinicians" with an impact factor of 144.8 showed that esophageal cancer, Gastric cancer and colorectal cancer are both among the top five types of cancer incidence among men and women in China, and colorectal cancer has shown an increasing trend in the past ten years. At present, the proportion of early diagnosis and treatment of digestive tract cancer is very low. Take colorectal cancer as an example: the proport...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886
Inventor 赵国栋
Owner SUZHOU VERSABIO TECH INC
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