TABLE 3.
Primers used in this study
| Target | Orientationa | Sequence (5′→3′) |
|---|---|---|
| Primers for site-directed mutagenesis | ||
| Pm25|E546A | F | GTGGGATGTGCTGAACGCGCCCATGAACGAGAAC |
| R | GTTCTCGTTCATGGGCGCGTTCAGCACATCCCAC | |
| Pm25|E663tga | F | GGCAAACTCATCAAGGTGACGTGACTCGATATTGCCATATCCAC |
| R | GTGGATATGGCAATATCGAGTCACGTCACCTTGATGAGTTTGCC | |
| Pm25|tga663A | F | GCAAACTCATCAAGGTGACGGCGCTCGATATTGCCATATCC |
| R | GGATATGGCAATATCGAGCGCCGTCACCTTGATGAGTTTGC | |
| Pm25|H634A | F | GCATCGGCACGCAAATGGCGCTCAACCTCAACTGG |
| R | CCAGTTGAGGTTGAGCGCCATTTGCGTGCCGATGC | |
| CBM4-1|Y213A | F | GGCTCGACCGGAAAAGCGGACGCACAACTCAACTCTCC |
| R | GGAGAGTTGAGTTGTGCGTCCGCTTTTCCGGTCGAGCC | |
| CBM4-1|Q216A | F | CGGAAAATACGACGCAGCGCTCAACTCTCCGGCG |
| R | CGCCGGAGAGTTGAGCGCTGCGTCGTATTTTCCG | |
| CBM4-1|N218A | F | TACGACGCACAACTCGCGTCTCCGGCGTTCCCG |
| R | CGGGAACGCCGGAGACGCGAGTTGTGCGTCGTA | |
| CBM4-1|Y257A | F | CGGAACTGGGTAATCAAGCGCCATGGGTGAACGGC |
| R | GCCGTTCACCCATGGCGCTTGATTACCCAGTTCCG | |
| CBM4-1|W259A | F | GGTAATCAATATCCAGCCGTGAACGGCGCAGAACTGGC |
| R | GCCAGTTCTGCGCCGTTCACGGCTGGATATTGATTACC | |
| CBM4-1|N261A | F | CAATATCCATGGGTGGCGGGCGCAGAACTGGCCG |
| R | CGGCCAGTTCTGCGCCCGCCACCCATGGATATTG | |
| CBM4-2|Y378A | F | GGTGCGGCTGGCAAAGCGGACCGCCAACTCAAC |
| R | GTTGAGTTGGCGGTCCGCTTTGCCAGCCGCACC | |
| CBM4-2|Q381A | F | GGCAAATACGACCGCGCGCTCAACAGCCCGGCATTTCC |
| R | GGAAATGCCGGGCTGTTGAGCGCGCGGTCGTATTTGCC | |
| CBM4-2|N383A | F | GCAAATACGACCGCCAACTCGCGAGCCCGGCATTTCCG |
| R | CGGAAATGCCGGGCTCGCGAGTTGGCGGTCGTATTTGC | |
| CBM4-2|Y422A | F | GAACTCGGCAACCAGGCGCCGTGGGGAAGCGGC |
| R | GCCGCTTCCCCACGGCGCCTGGTTGCCGAGTTC | |
| CBM4-2|W424A | F | GGCAACCAGTATCCGGCCGGAAGCGGCGGCGAG |
| R | CTCGCCGCCGCTTCCGGCCGGATACTGGTTGCC | |
| Primers for in-fusion cloning | ||
| Pm25_CBM4-1_L167-L320 | F | CAGCCATATGGCTAGCTTGTGGACGGAAGTTTCGAAG |
| R | GGTGGTGGTGCTCGATTAGAGGTCTGTGATCTCGACG | |
| Pm25_CBM4-2_D332-D484 | F | CAGCCATATGGCTAGCCTTGTGGACGGCAATTTCGAAGGCGGAG |
| R | GGTGGTGGTGCTCGAGTTATTAGAGGTCGGTGACCTCAACGGCGTC | |
| Pm25_CBM4-1 + 2_L167-D484 | F | GACGACGACGACAAGCTTGTGGACGGAAGTTTCGAAGAGGGGATG |
| R | GTGGTGGTGCTCGAGGTCGGTGACCTCAACGGCGTCGATC | |
| pET32a_linearization | F | CTCGAGCACCACCACCACCACCAC |
| R | CTTGTCGTCGTCGTCGCCAGAACCAGAACCGGCCAGGTTAG |
a
F, forward; R, reverse.