Construction of human epiplakin fragments and full length cDNA of human keratins
| cDNA fragment* (accession no.) . | Primers used in PCR† . | Corresponding amino acid region . |
|---|---|---|
| Epiplakin: | 5′-CTCTACcatatgCAGCTGGCAGGCCGGGG-3′ | 4527-5065 (L12-B13) |
| (AB051895) | 5′-GAGGAActcgagTCACTGTAGAGAGAGAGAAAGAAATAGG-3′ | |
| Epiplakin: | 5′-CTCTACcatatgCAGCTGGCAGGCCGGGG-3′ | 4527-4885 (L12) |
| 5′-GCTCATctcgagCTAGCGGCCGGGCTGGTCCTTGGC-3′ | ||
| Epiplakin: | 5′-GCCCcatatgCAGGAGAAGATGAGCATCTACCAGG-3′ | 4885-5065 (B13) |
| 5′-GAGGAActcgagTCACTGTAGAGAGAGAGAAAGAAATAGG-3′ | ||
| Keratin 5 | 5′-AGGAACAAGCCcatatgTCTCGCCAGTCAAGT-3′ | 1-590 (full length) |
| (NM_000424) | 5′-GTGACTTGCAGCAGGaagcttGCTCTTGAAGCTCT-3′ | |
| Keratin 14 | 5′-ACCTCCCTCCTCTGCcatatgACTACCTGCAGCCGCCAG-3′ | 1-472 (full length) |
| (NM_000526) | 5′-TGAGCGGGGCTGGGCAaagcttGTTCTTGGTGCGAAGGAC-3′ | |
| Keratin 8 | 5′-CTCCAGCCTCTcatatgTCCATCAGGGTGACCCAGAAGT-3′ | 1-483 (full length) |
| (NM_002273) | 5′-AGGGGCTGCCGCAgcggccgcCTTGGGCAGGACGTCAG-3′ | |
| Keratin 18 | 5′-TCTCTCTCCCCGGACcatatgAGCTTCACCACTCGCTCCACCT-3′ | 1-430 (full length) |
| (NM_000224) | 5′-AGGGTACCCTGCTTCTGCTaagcttATGCCTCAGAACTTTGGTG-3′ |
| cDNA fragment* (accession no.) . | Primers used in PCR† . | Corresponding amino acid region . |
|---|---|---|
| Epiplakin: | 5′-CTCTACcatatgCAGCTGGCAGGCCGGGG-3′ | 4527-5065 (L12-B13) |
| (AB051895) | 5′-GAGGAActcgagTCACTGTAGAGAGAGAGAAAGAAATAGG-3′ | |
| Epiplakin: | 5′-CTCTACcatatgCAGCTGGCAGGCCGGGG-3′ | 4527-4885 (L12) |
| 5′-GCTCATctcgagCTAGCGGCCGGGCTGGTCCTTGGC-3′ | ||
| Epiplakin: | 5′-GCCCcatatgCAGGAGAAGATGAGCATCTACCAGG-3′ | 4885-5065 (B13) |
| 5′-GAGGAActcgagTCACTGTAGAGAGAGAGAAAGAAATAGG-3′ | ||
| Keratin 5 | 5′-AGGAACAAGCCcatatgTCTCGCCAGTCAAGT-3′ | 1-590 (full length) |
| (NM_000424) | 5′-GTGACTTGCAGCAGGaagcttGCTCTTGAAGCTCT-3′ | |
| Keratin 14 | 5′-ACCTCCCTCCTCTGCcatatgACTACCTGCAGCCGCCAG-3′ | 1-472 (full length) |
| (NM_000526) | 5′-TGAGCGGGGCTGGGCAaagcttGTTCTTGGTGCGAAGGAC-3′ | |
| Keratin 8 | 5′-CTCCAGCCTCTcatatgTCCATCAGGGTGACCCAGAAGT-3′ | 1-483 (full length) |
| (NM_002273) | 5′-AGGGGCTGCCGCAgcggccgcCTTGGGCAGGACGTCAG-3′ | |
| Keratin 18 | 5′-TCTCTCTCCCCGGACcatatgAGCTTCACCACTCGCTCCACCT-3′ | 1-430 (full length) |
| (NM_000224) | 5′-AGGGTACCCTGCTTCTGCTaagcttATGCCTCAGAACTTTGGTG-3′ |
The epiplakin fragments were cloned into pGATEV expression plasmid. Full-length keratin cDNAs were cloned into pET23a plasmid and in frame with His-tag on the C-terminal
In each PCR reaction, the upper and lower primers are the sense and antisense oligonucleotides, respectively. The cloning linkers are lowercase. The stop codon of each epiplakin fragment is underlined