Method and system for constructing sequencing library on the basis of methylated DNA target region, and use thereof

A kit configured to construct a sequencing library based on a target region of a methylated DNA by a method for constructing a sequencing library based on a target region of a methylated DNA, the method comprising:

• step 1 of obtaining a transformed DNA sample with universal sequence based on a methylated DNA sample by ligating a universal sequence to at least one end of the methylated DNA sample and treating the methylated DNA sample with bisulfite;
• step 2 of performing, by using a first specific primer and a first universal primer, a first amplification on the transformed DNA sample with universal sequence to obtain a first amplification product, wherein the first specific primer is located upstream of the target region, the first universal primer is at least partially complementary to or at least partially overlaps the universal sequence, and the first universal primer is located downstream of the target region;
• and
• step 3 of performing, by using a second specific primer, a second universal primer and a tagged primer, a second amplification on the first amplification product to obtain a second amplification product and obtain the sequencing library, wherein the second specific primer, the second universal primer, and the tagged primer are set forth:
• i:
• the second specific primer is located downstream of the first specific primer and upstream of the target region, the second universal primer at least partially overlaps a sequence of the second specific primer, the tagged primer contains a tag sequence, and the tagged primer partially overlaps a sequence of the first universal primer;
• or
• ii:
• the second specific primer is located downstream of the target region, the second universal primer at least partially overlaps a sequence of the first specific primer, the tagged primer contains a tag sequence, and the tagged primer partially overlaps a sequence of the second specific primer,
• wherein the first specific primer and the second specific primer are designed for only one strand of the DNA sample,
• the kit comprising:
• a universal sequence to be ligated to at least one end of the methylated DNA sample, wherein the universal sequence is at least one selected from a sequencing adapter sequence or a modified sequencing adapter sequence, the sequencing adapter sequence being a known sequencing adapter sequence of a sequencing platform;
• a tagged primer set forth as SEQ ID NO:
• 23;
• a first universal primer set forth as SEQ ID NO:
• 21, wherein the first universal primer matches or overlaps at least a part of the universal sequence;
• a second universal primer set forth as SEQ ID NO:
• 22;
• a methylation detection reagent, wherein the methylation detection reagent is a bisulfite detection reagent;
• a first specific primer comprising a sequence set forth as SEQ ID NO:
• 1 to SEQ ID NO:
• 10;
• and
• a second specific primer comprising a sequence set forth as SEQ ID NO:
• 11 to SEQ ID NO:
• 20,
• wherein the first specific primer and the second specific primer are designed for only one strand of a DNA sample.