Composition for detecting esophageal cancer
A composition and technology for esophageal cancer, applied in the biological field, can solve the problems of high cost, inconvenient detection, low sensitivity and the like, and achieve the effects of effective detection and good detection sensitivity
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Embodiment 1
[0227] By analyzing the data of 233 cases of esophageal cancer tissues and 171 cases of normal esophagus tissues, the genome-wide methylation chip (Illumina's HumanMethylation450k chip) data, the inventors found that the methylation levels of MT1A gene and EPO gene in esophageal cancer tissues were significantly higher in normal esophageal tissue (analysis results such as figure 1 shown). Furthermore, the present inventors found that the two target genes were significantly different in esophageal cancer tissue and normal esophagus tissue by analyzing the probe sequences of MT1A gene and EPO gene on the genome-wide methylation chip and the corresponding methylation rate data. The sequence fragments with the most obvious difference in methylation were identified as the target sequences of the two target genes.
[0228] The target sequence of MT1A gene is shown in SEQ ID NO:1.
[0229] SEQ ID NO:1
[0230] CACCCAGGGGAGCTCAGTGGACTGTGCGCCTTGCCTTTCTGCTGCGCAAAGCCCAGTCCAGGTCATCACCT...
Embodiment 2
[0241] Step 1: Obtain the DNA of the biological sample to be analyzed. The source may be any suitable source, such as cell lines, histological sections, biopsies, paraffin-embedded tissues, body fluids, feces, urine, plasma, serum, whole blood, isolated blood cells, cells isolated from blood, and all possible combinations. DNA is then isolated from the sample by any standard means known in the art. Briefly, when DNA is enclosed in cell membranes, the biological sample must be disrupted and lysed by enzymatic, chemical or mechanical means. Proteins and other contaminants are subsequently removed, eg, by protein kinase K digestion. The DNA is then recovered from the solution. This can be achieved by various methods including salting out, organic extraction or binding the DNA to a solid support. The choice of method will be influenced by several factors, including time, cost, and the amount of DNA required. When the sample DNA is not encapsulated in cell membranes (eg, circu...
Embodiment 3
[0274] According to a specific embodiment of the present application, based on the average Ct value of the detection results of a certain number of esophageal cancer samples and normal samples, the Ct value of the target gene that can effectively distinguish esophageal cancer and normal samples is determined, that is, the critical value. The methylation status of at least one CpG dinucleotide of the target gene target sequence is determined by the cycle threshold Ct value of the polymerase chain reaction. By comparing the Ct value of the measured sample with the preset critical value, it is determined that the Whether the gene analysis is negative (normal) or positive (esophageal cancer).
[0275] This embodiment includes the following steps:
[0276] First, plasma samples from 20 patients with esophageal cancer and 22 normal individuals were obtained. All samples come from Boercheng Company. Then extract the free peripheral blood DNA of the test sample, and pretreat the DNA...
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