Gene marker for detecting esophageal cancer and use thereof and detection method
A gene marker and esophageal cancer technology, applied in the field of clinical molecular diagnosis of esophageal cancer, can solve the problems of lack of simple and effective
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Embodiment 1
[0033] Example 1: Screening of esophageal cancer gene markers:
[0034] (1) Extraction of plasma DNA: 10 ng of plasma DNA was extracted from samples from 20 patients with esophageal cancer and 20 normal subjects. This step can be carried out using any methods and reagents suitable for extracting plasma DNA well known to those skilled in the art.
[0035] (2) End-fill the plasma DNA, overhang A and connect with the sequencing adapter:
[0036] Prepare a reaction mixture containing 50uL plasma DNA, 7uL End Repair & A-Tailing Buffer and 3uL End Repair & A-Tailing Enzyme mix according to the Kapa Hyper Perp Kit instructions (the total volume is 60uL), incubate at 20°C for 30min, then incubate at 65°C ℃ warm bath for 30min. Prepare the following ligation reaction mixture in a 1.5mL low adsorption EP tube: 5uL Nuclease free water, 30uL Ligation Buffer and 10 uL DNA Ligase. 5uL of sequencing adapters were added to 45uL of the ligation reaction mixture, mixed, heated at 20°C for 20...
Embodiment 2
[0052] Example 2: Effectiveness of esophageal cancer gene markers:
[0053] This example is used to verify the effectiveness of the esophageal cancer gene markers of the present invention in detecting esophageal cancer. According to the method of Example 1, the 5-hmC content of the 10 esophageal cancer gene markers of the present invention in the first batch of 82 samples (41 cases of esophageal cancer and 107 cases of healthy controls) was determined. The standardized 5-hmC content of each gene marker is multiplied by the corresponding weighting coefficient of the marker in Example 1 to obtain the predictor t of the gene marker, and then the predictor t of each gene marker is added to obtain The total predictor T, and then the total predictor T is obtained by Logistic transformation according to the following formula to obtain the score P: P=1 / (1+e -T ), if P>0.5, the subject sample has esophageal cancer; if P≤0.5, the subject sample is normal. figure 1 The results of disti...
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