Primers used to construct the PS mutants F1FII-GlaPS-PS, F2FII-GlaPS-PS, and F12PS-GlaFII-PS
Primers . | Sequences . |
|---|---|
| EcoFII, s | 5′-CTCACTATAGGCTAGCCTCGAgaattcTTCGAAATGGCGCACGTCCGAGGCTTGCAG-3′ |
| EcoPS, s | 5′-CTCACTATAGGCTAGCCTCGAgaattcTTCGAAATGAGGGTCCTGGGTGGGCGCTGCGGG-3′ |
| F1FII-1, a | 5′-GTATTTTGGATAAAACACATCCGTAGCCGTGTCGGATTCAAAGACCTCCCTGGCTTCTTC-3′ |
| F1FII-2, s | 5′-TCCGACACGGCTACGGATGTGTTTTATCCAAAATACTTAGTTTGTCTT-3′ |
| F2FII-1, a | 5′-TGTGTATTTTGGCCAAAAATAATCCGTTTCCGGGGAATTTTCAAAGACCTCCCTGGCTTC-3′ |
| F2FII-2, s | 5′-TCCCCGGAAACGGATTATTTTTGGCCAAAATACACAGTTTGTCTTCGCTCTTTTCAAACT-3′ |
| F2FII-3, a | 5′-TTCAAAGACCTCGAAGGCTTCTTCGTAGCTGCACGTTTCTTCGATGCATTCTCTTTCAAG-3′ |
| F2FII-4, s | 5′-ACGTGCAGCTACGAAGAAGCCTTCGAGGTCTTTGAAAATTCCCCGGAA-3′ |
| F12PS-1, a | 5′-TAAGTACTTCGGATAGAAATAATCCGTTTCCGGGTCATTCTCCAGAGCCTCGAAGGCCTCCTC-3′ |
| F12PS-2, s | 5′-AATGACCCGGAAACGGATTATTTCTATCCGAAGTACTTAGTTTGTCTTCGCTCTTTTCAAACT-3′ |
| F12PS-3, a | 5′-CTCCAGAGCCTCCCTGGCCTCCTCTTTATTGCACAGCTCCTCCACGCACTCTCGCTCTAG-3′ |
| F12PS-4, s | 5′-CTGTGCAATAAAGAGGAGGCCAGGGAGGCTCTGGAGAATGACCCGGAA-3′ |
| Afl, a | 5′-AGTGTAACCTCCAGGGTAATTGACACAAAGCTGAGCACacatgtTCTCAGAGCATT-3′ |
Primers . | Sequences . |
|---|---|
| EcoFII, s | 5′-CTCACTATAGGCTAGCCTCGAgaattcTTCGAAATGGCGCACGTCCGAGGCTTGCAG-3′ |
| EcoPS, s | 5′-CTCACTATAGGCTAGCCTCGAgaattcTTCGAAATGAGGGTCCTGGGTGGGCGCTGCGGG-3′ |
| F1FII-1, a | 5′-GTATTTTGGATAAAACACATCCGTAGCCGTGTCGGATTCAAAGACCTCCCTGGCTTCTTC-3′ |
| F1FII-2, s | 5′-TCCGACACGGCTACGGATGTGTTTTATCCAAAATACTTAGTTTGTCTT-3′ |
| F2FII-1, a | 5′-TGTGTATTTTGGCCAAAAATAATCCGTTTCCGGGGAATTTTCAAAGACCTCCCTGGCTTC-3′ |
| F2FII-2, s | 5′-TCCCCGGAAACGGATTATTTTTGGCCAAAATACACAGTTTGTCTTCGCTCTTTTCAAACT-3′ |
| F2FII-3, a | 5′-TTCAAAGACCTCGAAGGCTTCTTCGTAGCTGCACGTTTCTTCGATGCATTCTCTTTCAAG-3′ |
| F2FII-4, s | 5′-ACGTGCAGCTACGAAGAAGCCTTCGAGGTCTTTGAAAATTCCCCGGAA-3′ |
| F12PS-1, a | 5′-TAAGTACTTCGGATAGAAATAATCCGTTTCCGGGTCATTCTCCAGAGCCTCGAAGGCCTCCTC-3′ |
| F12PS-2, s | 5′-AATGACCCGGAAACGGATTATTTCTATCCGAAGTACTTAGTTTGTCTTCGCTCTTTTCAAACT-3′ |
| F12PS-3, a | 5′-CTCCAGAGCCTCCCTGGCCTCCTCTTTATTGCACAGCTCCTCCACGCACTCTCGCTCTAG-3′ |
| F12PS-4, s | 5′-CTGTGCAATAAAGAGGAGGCCAGGGAGGCTCTGGAGAATGACCCGGAA-3′ |
| Afl, a | 5′-AGTGTAACCTCCAGGGTAATTGACACAAAGCTGAGCACacatgtTCTCAGAGCATT-3′ |
Sequences complementary to the prothrombin cDNA are shown in bold, and sequences complementary to the PS cDNA are shown in italics. For primers EcoFII and EcoPS, sequences complementary to the pCI-neo vector, upstream of the initial codon of the PS cDNA, are in uppercase letters. Primers EcoFII and EcoPS contain an EcoRI restriction site (lowercase letters), and primer Afl contains an Af/III restriction site (lowercase letters) for cloning purposes. For primer pairs F1FII-1/F1FII-2, F2FII-1/F2FII-2, F2FII-3/F2FII-4, F12PS-1/F12PS-2, and F12PS-3/F12PS-4, the sequence underlined in each antisense primer is complementary to the sequence underlined in each sense primer.
Abbreviations are shown in Table 1.