Table 1.

Oligonucleotide sequences and specific PCR conditions for the real-time quantitative PCR and digital PCR assays.

AssayOligonucleotideSequences (5′ to 3′)Specific thermal cycling conditions
HLCS (real-time quantitative PCR and digital PCR)Forward primerCCGTGTGGCCAGAGGTG50 Cycles of 95 °C for 30 s and 60 °C for 1 min
Reverse primerAAAGGGCCAGGTCGGGA
Hydrolysis probeFAM-AGGATTTGGGGCTGCGC (MGB)1
RASSF1A (digital PCR)Forward primerAGCTGGCACCCGCTGG50 Cycles of 95 °C for 30 s and 60 °C for 1 min
Reverse primerGTGTGGGGTTGCACGCG
Hydrolysis probeVIC-ACCCGGCTGGAGCGT (MGB)2
ZFX/ZFY (digital PCR)Forward primerCAAGTGCTGGACTCAGATGTAACTG50 Cycles of 95 °C for 15 s and 57 °C for 1 min
Reverse primerTGAAGTAATGTCAGAAGCTAAAACATCA
Hydrolysis probe_ZFYFAM-TCTTTACCACACTGCAC (MGB)
Hydrolysis probe_ZFXVIC-TCTTTAGCACATTGCA (MGB)