PCR primer and probe sequences
Oligo name . | Sequence (5′-3′) . |
|---|---|
| Taqman primers and probes | |
| 5′ RE-IIBP | TGATATGTCTGGGATTTGCTTCAG |
| 3′ RE-IIBP | GCCCCCAAAAACATCAACCT |
| RE-IIBP probe | CGTGGCTCTATCCATAC |
| 5′ Exon 4a | CACTCCTTCACAGCTATACCAAACTT |
| 3′ Exon 4a | GGGAGGAGAGAGGAAGGCA |
| Exon 4a probe | TTGTTTTTAAGTGTTCAAACTTC |
| 5′ MMSET I | CATCTCCTTCTGCATCCTTAACTGA |
| 3′ MMSET I | AGCAGCTGGGTTCAAATCCAA |
| MMSET I probe | CTCCCACAAAAGCTC |
| 5′ MMSET II | TCATTTGGGTGAAACTTGGGAACT |
| 3′ MMSET II | TCCAATCTCGTGCTTCATTTTCTGA |
| MMSET II probe | CCGGCCACCATCTGT |
| Cloning primers | |
| 5′ MMSET (NdeI) | gcatatgCTGGATGGAATTTAGC |
| 5′ MB4-2 (NdeI) | gcatatgATTCCAGCTAAGAAAGAG |
| 5′ MB4-3 (NdeI) | gcatatgGTCAGAAAAAGAGTGCAC |
| 5′ RE-IIBP (NdeI) | gcatatgTAAAATGATGCGGTG |
| 3′ MMSET I (XhoI) | gctcgagCTAAGTGCAGTACAGAGC |
| 3′ MMSET II (XhoI) | gctcgagCTATTTGCCCTCTGTC |
| 3′ MMSET Exon 4a (XhoI) | gctcgagTTAATCTTTCAGTACAATTTG |
| 3′ MMSET I (Δstop-XhoI) | gctcgagGTGCAGTACAGAGCAGC |
| 3′ MMSET II (Δstop-XhoI) | gctcgagTATTTGCCCTCTGTGAC |
| 3′ Exon 4a (Δstop-XhoI) | gctcgagTCTTTCAGTACAATTTGAC |
| 5′ MMSET (XhoI) | gCGActcgagGCTGGATGGAATTTAG |
| 3′ Exon 4a (Δstop-EcoRI) | gAGCgaattcAGTgcATCTTTCAGTACAAT |
Oligo name . | Sequence (5′-3′) . |
|---|---|
| Taqman primers and probes | |
| 5′ RE-IIBP | TGATATGTCTGGGATTTGCTTCAG |
| 3′ RE-IIBP | GCCCCCAAAAACATCAACCT |
| RE-IIBP probe | CGTGGCTCTATCCATAC |
| 5′ Exon 4a | CACTCCTTCACAGCTATACCAAACTT |
| 3′ Exon 4a | GGGAGGAGAGAGGAAGGCA |
| Exon 4a probe | TTGTTTTTAAGTGTTCAAACTTC |
| 5′ MMSET I | CATCTCCTTCTGCATCCTTAACTGA |
| 3′ MMSET I | AGCAGCTGGGTTCAAATCCAA |
| MMSET I probe | CTCCCACAAAAGCTC |
| 5′ MMSET II | TCATTTGGGTGAAACTTGGGAACT |
| 3′ MMSET II | TCCAATCTCGTGCTTCATTTTCTGA |
| MMSET II probe | CCGGCCACCATCTGT |
| Cloning primers | |
| 5′ MMSET (NdeI) | gcatatgCTGGATGGAATTTAGC |
| 5′ MB4-2 (NdeI) | gcatatgATTCCAGCTAAGAAAGAG |
| 5′ MB4-3 (NdeI) | gcatatgGTCAGAAAAAGAGTGCAC |
| 5′ RE-IIBP (NdeI) | gcatatgTAAAATGATGCGGTG |
| 3′ MMSET I (XhoI) | gctcgagCTAAGTGCAGTACAGAGC |
| 3′ MMSET II (XhoI) | gctcgagCTATTTGCCCTCTGTC |
| 3′ MMSET Exon 4a (XhoI) | gctcgagTTAATCTTTCAGTACAATTTG |
| 3′ MMSET I (Δstop-XhoI) | gctcgagGTGCAGTACAGAGCAGC |
| 3′ MMSET II (Δstop-XhoI) | gctcgagTATTTGCCCTCTGTGAC |
| 3′ Exon 4a (Δstop-XhoI) | gctcgagTCTTTCAGTACAATTTGAC |
| 5′ MMSET (XhoI) | gCGActcgagGCTGGATGGAATTTAG |
| 3′ Exon 4a (Δstop-EcoRI) | gAGCgaattcAGTgcATCTTTCAGTACAAT |
Taqman primer and probe pairs were designed by Applied Biosystems using the Assays-by-Design service. All probes contain the 5′ reporter dye 6-FAM (carboxyfluorescein) and the 3′ minor groove binder/nonfluorescence quencher (MGB/NFQ). The start and stop codons in the cloning primers are underlined; mutated and restriction endonuclease target sequences are indicated by lowercase letters. Any mutations incorporated within coding regions are silent mutations that maintain the proper amino acid sequence.