Table 1.
Primers utilized in this study.
| Reaction | Name | Primer Sequence (5′ to 3′) | Tm (°C) | Product Length (bp) | Amplified Region | Accession numbers |
|---|---|---|---|---|---|---|
| RT-PCR | β-actin | F: CACCAACTGGGACGACAT | 60.0 | 202 | 320–521 | AY141970.1 |
| R: ATACAGGGACAGCACAGC | ||||||
| SIX 1 | F: GCCAAGGAAAGGGAGAACA | 60.0 | 127 | 866–992 | XM_588692.7 | |
| R: GACTCTGGGGAGGTGAGAACT | ||||||
| CDS | SIX1 CDSF/R | F: ATGTCGATGCTGCCATCGTTC | 62.0 | 855 | 317–1171 | XM_588692.7 |
| R: TTAGGACCCCAAGTCCACCA | ||||||
| Promoter cloning | SIX1-PF/PR | F: ATGAGCACTAAAATAAGACCATGAGGG | 65.5 | 2171 | −2000/+170 | NC_007308.6 |
| R: CTTGAAGACTGGAGGCCGAGTA | ||||||
| SIX1-P1 | F: CGGGGTACCATGAGCACTAAAATAAGACC | 64.0 | 2171 | −2000/+170 | ||
| SIX1-P2 | F: CGGGGTACCATCTCCTGCCAGTCTCAGCTAC | 61.5 | 1471 | −1300/+170 | ||
| SIX1-P3 | F: CGGGGTACCACTATGCTCTACACAATTCTTCTC | 63.5 | 860 | −689/+170 | ||
| SIX1-P4 | F: CGGGGTACCCTGGCCAGCCCCGGATAGGCG | 65.0 | 211 | −40/+170 | ||
| SIX1-R | R: GGAAGATCTCTTGAAGACTGGAGGCCGAGTA | |||||
| Site-mut and EMSA | MyoD forward | GGAAGCGCTATACAGCTGGGAGTATTTGTGCGCT | −630/−596 | |||
| MyoD reverse | AGCGCACAAATACTCCCAGCTGTATAGCGCTTCC | |||||
| mMyoD forward | GGAAGCGCTATACAAAAAGGAGTATTTGTGCGCT | −630/−596 | ||||
| mMyoD reverse | AGCGCACAAATACTCCTTTTTGTATAGCGCTTCC | |||||
| PAX7 forward | CATAAGCTATTCATCAATCCATTACTTATTCATT | −455/−431 | ||||
| PAX7 reverse | AATGAATAAGTAATGGATTGATGAATAGCTTATG | |||||
| mPAX7 forward | CATAAGCTATTCATCTTTTCATTACTTATTCATT | −455/−431 | ||||
| mPAX7 reverse | AATGAATAAGTAATGAAAAGATGAATAGCTTATG | |||||
| CREB forward | CACGTTGCAAGGTCCTGACGCGCTCACCCACTCC | −259/−225 | ||||
| CREB reverse | GGAGTGGGTGAGCGCGTCAGGACCTTGCAACGTG | |||||
| mCREB forward | CACGTTGCAAGGTCCTTTTGCGCTCACCCACTCC | −259/−225 | ||||
| mCREB reverse | GGAGTGGGTGAGCGCAAAAGGACCTTGCAACGTG | |||||
| MEF3 forward | CACAGCAGAGGGGGCTCACGTTGCAAGGTCCTGA | −274/−240 | ||||
| MEF3 reverse | TCAGGACCTTGCAACGTGAGCCCCCTCTGCTGTG | |||||
| mMEF3 forward | CACAGCAGAGGGGGCAAAAGTTGCAAGGTCCTGA | −274/−240 | ||||
| mMEF3 reverse | TCAGGACCTTGCAACTTTTGCCCCCTCTGCTGTG | |||||
| ChIP | ChIP-MyoD | F: GCTCAGCAACTATGCTCTACA | 60 | 112 | −697/−586 | |
| R: CGCCCTCCAAAGCGCACAAAT | ||||||
| ChIP-PAX7 | F: GAGGACTGGTCTACGCTTGTAT | 60 | 124 | −491/−367 | ||
| R: CTTTGCAGACTTGCCATTTCC | ||||||
| ChIP-CREB | F: GAGCCATTGATTTGTGCAAAGATG | 60 | 165 | −319/−154 | ||
| R: GCGAGTGGTGAGAAGCGAAGTG | ||||||
| ChIP-MyoG | F: GAGCCATTGATTTGTGCAAAGATG | 60 | 165 | −319/−154 | ||
| R: GCGAGTGGTGAGAAGCGAAGTG | ||||||
| ChIP-control | F: GTTTTGTTTACCACTAGCTTTTC | 60 | 134 | SIX1 exon 2 | ||
| R: ATCCTTGTAGGAGTTCCCTTT | ||||||
| siRNA | siMyoD | CCAAUGCGAUUUAUCAGGUGCUUUGTT | ||||
| siPAX7 | GGUAACAUCCCAGCUUUACTT | |||||
| siCREB | AAUACAGCUGGCUAACAAUGGTT | |||||
| siMyoG | AACUACCUUCCUGUCCACCTT | |||||
| siRNA-NC | UUCUCCGAACGUGUCACGUTT | |||||
| Overexpression | MyoG-CDSF/R | F: CCCAAGCTTATGGAGCTGTATGAGACCTCTC | 62.5 | 675 | NM_001111325.1 | |
| R: CTAGTCTAGATCAGTTTGGTATGGTTTCATCTG |
The italicized letters of the forward (F) and reverse (R) primers for promoter cloning and overexpression items indicate the enzyme cutting sites of promoter cloning (KpnI and BglII) and overexpression items (HindIII and Xbal), respectively. The underlined bases are core putative transcription factor-binding sites.