TABLE 1.
Oligonucleotide primers for P mutations
| Primer group and mutant | Primer sequencea | Enzyme |
|---|---|---|
| Mutagenic primer group | ||
| P408/9 | (+)-CGGATATCTACGCGGCATTCTCTGAG | EcoRV |
| P1 | (+)-CTCTGCGTATCAGGCCGCGCAGAACTC | BsoFI |
| S419A | (+)-GAACAGAACGCATTGCTCATGTCCAACC | NlaIII |
| S426A | (+)-CCAACCTAGCTACGCTGCATATCATCAC | BsoFI |
| P2 | (+)-CATCACAGCTGCAGGTGGCGCGACTGACAAC | BsoFI |
| P3 | (+)-CCTCCGTTTTTGCAGCATCAGCGGCCACCGCGACTAAGGC | BsoFI |
| K453A | (+)-CCTCCGTTTTTGCAGCATCAAAAGAGAACAAGACTAAGGC | BsoFI |
| P455/6 | (+)-CCTCCGTTTTTGCAAAATCAGCAGCGAACAAGACTAAGGC | BsoFI |
| P4 | (+)-CAAGACTGCGGCTACCGCGTTTGCCCCATC | BsoFI |
| P5 | (+)-CCATCTATGGCCACCCTAGCCGCGATGAAGTACAAACCG | BsoFI |
| L421A | (+)-CAGAACTCATTGGCCATGTCCAACCTATC | NlaIII |
| L425A | (+)-GCTGATGTCCAACGCTAGCACACTTCATATCATCAC | NheI |
| L428A | (+)-GATGTCCAACCTAAGTACTGCTCATATCATCACAG | ScaI |
| I430A | (+)-CAACCTATCTACACTGCACGCCATCACAGATAGAG | BsgI |
| G436A | (+)-CACAGATAGAGGCGCCAAGACTGACAAC | HaeII |
| Outside primer group | ||
| Primer 1 | (+)-ATCTACACAGGATGAGC | |
| Primer 2 | (−)-GATCTCGAGCCCGGGATCTAGTTG | XhoI |
a
The oligonucleotide sequences are written 5′ to 3′. The plus symbol refers to the messenger sense (+) of the oligonucleotide. The underlined sequence indicates the introduced silent restriction enzyme site shown on the right.