Esophageal cancer gene methylation level detection reagent and application thereof
A methylation and reagent technology, applied in the field of esophageal cancer gene methylation level detection reagents, can solve the problems of low cfDNA content, lack, extraction difficulties, etc., and achieve high sensitivity and specificity
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Embodiment 1
[0047] This embodiment provides reagents for the diagnosis or auxiliary diagnosis of esophageal cancer or precancerous lesions, which include nucleic acid combination 1.
[0048] Nucleic acid combination 1 includes the nucleotides shown in SEQ ID NO.1-3. This nucleic acid combination 1 can detect the methylation level of the Chr17: 74923881-74924015 region positive strand (target region 1) on the OTOP2 gene.
[0049] The upstream primer sequence for region 1 methylation-specific PCR is (5’-3’):
[0050] TAGGAGTATAGGATTTAGGGTGCGT (SEQ ID NO. 1);
[0051] The downstream primer sequences for region 1 methylation-specific PCR are (5'-3'):
[0052] TAATACCCACCGCGCCCTAAC (SEQ ID NO. 2);
[0053] The probe sequence for region 1 methylation-specific PCR is (5'-3'):
[0054] TTGGAGGGCGTTAGGGTGTTCGT (SEQ ID NO. 3).
Embodiment 2
[0056] This embodiment provides reagents for the diagnosis or auxiliary diagnosis of esophageal cancer or precancerous lesions, which include nucleic acid combination 2.
[0057] Nucleic acid combination 2 includes the nucleotides shown in SEQ ID NO.4-6. This nucleic acid combination 2 can detect the methylation level of the Chr17: 74924159-74924335 region positive strand (target region 2) on the OTOP2 gene.
[0058] The upstream primer sequences for region 2 methylation-specific PCR are (5’-3’):
[0059] TTGGAGAAAGATGGATTATGTGAGC (SEQ ID NO. 4);
[0060] The downstream primer sequences for region 2 methylation-specific PCR are (5'-3'):
[0061] ACCTAACTAAACTTAAAACCGACGC (SEQ ID NO. 5);
[0062] The probe sequence for region 2 methylation-specific PCR is (5'-3'):
[0063] AAAGCGTTTGTTTATTTCGGCGTGG (SEQ ID NO. 6).
Embodiment 3
[0065] This embodiment provides reagents for the diagnosis or auxiliary diagnosis of esophageal cancer or precancerous lesions, which include nucleic acid combination 3.
[0066] Nucleic acid combination 3 includes the nucleotides shown in SEQ ID NO.7-9. This nucleic acid combination 3 can detect the methylation level of the Chr17: 74924422-74924581 region positive strand (target region 3) on the OTOP2 gene.
[0067] The upstream primer sequences for region 3 methylation-specific PCR are (5'-3'):
[0068] TTATTTAGCGAGAGGGGTAGGTTTC (SEQ ID NO. 7);
[0069] The downstream primer sequences for region 3 methylation-specific PCR are (5'-3'):
[0070] CAAAACTCCAAATCAACCCTAACGA (SEQ ID NO. 8);
[0071] The probe sequence for region 3 methylation-specific PCR is (5'-3'):
[0072] CGAAGTTTTAATTTTGCGGGTTAGG (SEQ ID NO. 9).
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