Table 2.
Primer Sequences and Conditions for qRT-PCR, Bisulfite sequencing and MSP Analysis
| Gene symbol | RefSeq mRNA | Primer Forward (5'-->3') | Primer Reverse (5'-->3') | Size (bp) | AT† (°C) |
| Quantitative Real Time PCR (qPCR) assay | |||||
| ABCA8 | NM_007168 | CCATCATGGTATCTGGGAGGTT | GCAGGTAATCTTTGCCAAATTTG | 77 | 60 |
| AQP8 | AB013456 | CACGGGCTGGCTTTGG | CCAGTACGGGAGGAGCATCA | 128 | 60 |
| CLCA4 | NM_012128 | CAAAATGGCCTATCTCAGTATTCCA | TCGCTTTGGCTTGAAGATTGT | 68 | 60 |
| HPGD1 | NM_000860 | GCATGGCATAGTTGGATTCACA | AAGCCTGGACAAATGGCATT | 83 | 60 |
| PRDX6 | NM_004905 | GCCCTTTCAATAGACAGTGTTGAG | ATCGATGATGGGAAAAGGTAACTT | 104 | 60 |
| SLC26A3 | NM_000111 | ATCGTTGGAACTGATGATGACTTC | CAGCATCATGGATTGTTAAGAAAAA | 90 | 60 |
| STX12 | NM_177424 | AAGAAAGAGAAACGGCAATTCG | TCATGGCCAAATCTTTAAATATCTGA | 75 | 60 |
| CSE1L | NM_001316 | CGGTTCAAACACAATAGCAAGTG | GGATGCAATCAGCTTCTGAAAGA | 78 | 60 |
| HSPH1 | NM_00664 | AACAAAATCCCAGATGCTGACA | ACCTTTATTTTGGGCTTTTTAGCTT | 79 | 60 |
| NEBL1 | HSY1624 | ATCCATGAGATCAATGCAGCAT | CATCCTGGGCACTGTAATCGT | 73 | 60 |
| RFC3 | NM_002915 | CTGAGGGAGACTGCAAATGCT | ACAGCCTTCCACGAACTTCAA | 72 | 60 |
| SLC12A2 | NM_001046 | AAGGAACATTCAAGCACAGCTAATATT | TGCCATGTAGAGAGCACTAGACACA | 86 | 60 |
| SOX9 | NM_000346 | ACGCCGAGCTCAGCAAGA | CACGAAGGGCCGCTTCT | 70 | 60 |
| GTF2IRD1 | ENST00000265755 | AATCTGCAATGATGCCAAGGT | CCGAGACCCGCTTTCTCTT | 70 | 60 |
| MXI1 | NM_005962 | CGGCACACAACACTTGGTTT | GGCTTTTTCTTTCAGCTTCTTCA | 75 | 60 |
| NR3C2 | NM_000901 | TTCATTCTCAGTACCAATAAAGCAAGA | GGTTTACTGTTGGATTCCCTTTAAAA | 82 | 60 |
| SGK1 | NM_005627 | AGGAGCCTGAGCTTATGAATGC | GACGACGGGCCAAGGTT | 75 | 60 |
| NDRG2 | NM_201535 | CTGACCGAGGCCTTCAAGTACT | GGCGAGTCATGCAGGATGA | 66 | 60 |
| TPX2 | NM_012112 | AGCCCTTTGTTCCCAAGAAAG | CCAGCTGAAAAGGTTCCTGAA | 80 | 60 |
| UBE2C | NM_181799 | TGGAGCTTACTCTGCAACTGTTTC | CCAAATGCCAGAACCCAACATTGATAGTCC | 74 | 60 |
| CCNB1 | NM_031966 | CCTGGCTAAGAATGTAGTCATGGTAA | GCATGCTTCGATGTGGCATA | 83 | 60 |
| SCNN1B | ENST00000343070 | CATTGAAGAATCAGCAGCCAATA | CCCATCCAGAAGCCAAACTG | 74 | 60 |
| FOXM1 | U74613 | AGCAAGCGAGTCCGCATT | CTGCAGAAGAAAGAGGAGCTATCC | 68 | 60 |
| SGK2 | NM_016276 | ACATCATTTACAGGGATCTGAAACC | TCCTTGCAGAGGCCAAAATC | 87 | 60 |
| Bisulfite Sequencing Analysis (BSA) | |||||
| NDRG2 | TTTTCGAGGGGTATAAGGAGAGTTTATTTT | CCAAAAACTCTAACTCCTAAATAAACA | 320 | 53 | |
| CSE1L | GTTTGGAATTTTAGTATTTTGGGAG | CTCTAACCATACCAACAAACTTCAC | 285 | 60 | |
| HSP1 | GGAGAGGGTTTGGGTATGTAA | CAAAAAAATAAAATAAACCTAAAAAAC | 194 | 56 | |
| PRDX6 | TATTTTTTTGTAGGGAGTTGGT | TAACATCCTTCAAACACTATAAACC | 279 | 56 | |
| SOX9 | TTTTTATTGATTTTTTTTGTAAAAG | ATACCAAAATTTTAATACCTTCTCC | 388 | 53 | |
| Methylation-Specific PCR (MSP) assay? | |||||
| NDRG2_UnM | AGAGGTATTAGGATTTTGGGTATGA | CCACTAAAAAAACAAAAATCTCACC | 125 | 55 | |
| NDRG2_M | AGAGGTATTAGGATTTTGGGTACG | GCTAAAAAAACGAAAATCTCGC | 123 | 55 | |
| NDRG2_UnM_2 | GGTAAATTTATTTGGGTATTGA | CAAAAACAAAATTAACCCTACAAA | 210 | 54 | |
| NDRG2_M_2 | TAGTGGTAAATTTATTCGGGTATCG | CAAAAACGAAATTAACCCTACGA | 214 | 62 | |
| p16-UnM | TTATTAGAGGGTGGGGTGGATTGT | CAACCCCAAACCCACAACCATAA | 151 | 65 | |
| p16-M | TTATTAGAGGGTGGGGCGGATCGC | GACCCCCGAACCGCGAACCGTAA | 150 | 68 | |
| APC-UnM | GTGTTTTATTGTGGAGTGTGGGTT | CCAATCAACAAACTCCCAACAA | 108 | 63 | |
| APC-M | TATTGCGGAGTGCGGGTC | ACCACCTCATCATAACTACCCACA | 98 | 63 | |
| MLH1-UnM | TTTTGATGTAGATGTTTTATTAGGTTGT | ACCACCTCATCATAACTACCCACA | 124 | 60 | |
| MLH1-M | ACGTAGACGTTTTATTAGGGTCGC | CCTCATCGTAACTACCCGCG | 115 | 60 |
?UnM = unmethylated sequence; M = methylated sequence.
† AT = annealing temperature.