Patent ID: 9605305
Date: 2017-03-28
CPC Classifications: C12Q

Claim:
1. A method for reducing primer-dimer amplification in a multiplex polymerase chain reaction (PCR), comprising the steps of: (a) obtaining a first nucleic acid sequence comprising a first tag (t1) and a first forward primer (F1) complementary to a first target nucleic acid fragment, (b) obtaining a second nucleic acid sequence comprising a second tag (t2) and a first reverse primer (R1) complementary to the first target nucleic acid fragment, (c) obtaining a third nucleic acid sequence comprising a third tag (t3) and a second forward primer (F2) complementary to a second target nucleic acid fragment, (d) obtaining a fourth nucleic acid sequence comprising the first tag (t1), a second reverse primer (R2) complementary to the second nucleic acid fragment, and a 5′-end partial sequence (F1^) or a full sequence of the first forward primer (F1) in between the first tag (t1) and the second reverse primer (R2), wherein the first forward primer (F1) and the second reverse primer (R2) have a complementary region at their 3′ends, F1^ has 3-30 nucleotides or 40-90% of the 5′-end partial F1 sequence, (e) mixing the first and the second target nucleic acid fragments, the first, the second, the third, and the fourth nucleic acid sequences, and an effective amount of reagents necessary for performing a polymerase chain reaction (PCR); and (f) performing PCR.