High-throughput sequencing method for methylated CpG island in trace DNA

A method for high-throughput sequencing of methylated CpG island in trace DNA, comprising the following steps:

• 1) performing a bisulfite treatment on a trace amount of DNA sample to obtain a single-stranded DNA sample converted with bisulfite;
• 2) adding the sample obtained in step 1) to a PCR system containing a primer A and linearly amplifying the processed sample obtained in step 1) in a PCR instrument;
• 3) adding an exonuclease having single-stranded DNA cleavage activity to a PCR product obtained in step 2) and reacting therewith at 37° C. for 30 minutes, and then increasing the temperature to inactivate the exonuclease;
• 4) adding an enzyme digestion product in step 3) to a PCR system containing a primer B for linear amplification;
• 5) then adding a PCR product system in step 4) to a PCR system containing a corresponding adapter primer C and an adapter primer D for PCR amplification;
• 6) size-selectively purificating a PCR product in step 5) to obtain a DNA library with a specific length, and performing high-throughput sequencing after passing quality control required by a sequencer,
• wherein the first base at the beginning of the 3′ portion of the primer B is D, and the immediate second base and third base constitute one CpG, while the 4th-9th bases contain one base G and the other bases are D, wherein D is A or T or G.