Table 3.
Oligonucleotides used in this study.
| Primer name | Sequence |
|---|---|
| A. FOR CONSTRUCTION OF pTA131-HfxMCM-HXba | |
| HfxMCM-5H | 5'-GTGTGTGTGTAAGCTTCCTCCGCGAGGCGACGGA-3' |
| HfxMCM-3X | 5'-GGTGGTGGTGTCTAGACATGGCAATCTTCTCCTG-3' |
| B. FOR CONSTRUCTION OF β7-β8 β-HAIRPIN MUTANTS BY OEM | |
| HFXMCM-rM1-F HFXMCM-rM1-R | 5'-AAACTGCGCGTCGCCGAGTCCCCCGAGGGC-3' 5'-GCCCTCGGGGGACTCGGCGACGCGCAGTTT-3' |
| HFXMCM-rM2-F HFXMCM-rM2-R | 5'-AAACTGCGCGTCCAGGCCTCCCCCGAGGGCCTGCG-3' 5'-CGCAGGCCCTCGGGGGAGGCCTGGACGCGCAGTTT-3' |
| HFXMCM-rM3-F HFXMCM-rM3-R | 5'-CAGGAGTCCCCCGCCGGCCTGCGCGGGGGC-3' 5'-GCCCCCGCGCAGGCCGGCGGGGGACTCCTG-3' |
| HFXMCM-rM4-F HFXMCM-rM4-R | 5'-CCCGAGGGCCTGGCGGGGGGCGAGACGCCG-3' 5'-CGGCGTCTCGCCCCCCGCCAGGCCCTCGGG-3' |
| HFXMCM-rM5-F HFXMCM-rM5-R | 5'-CTGCGCGGGGGCGCCACGCCGCAGAGCATC-3' 5'-GATGCTCTGCGGCGTGGCGCCCCCGCGCAG-3' |
| HFXMCM-rM6-F HFXMCM-rM6-R | 5'-GGCGAGACGCCGGCCAGCATCGACATCAAC-3' 5'-GTTGATGTCGATGCTGGCCGGCGTCTCGCC-3' |
| HFXMCM-rD1-F HFXMCM-rD1-R | 5'-CAGGAGTCCCCCGAG-GGCGAGACGCCGCAG-3' 5'-CTGCGGCGTCTCGCC-CTCGGGGGACTCCTG-3' |
| HFXMCM-rD2-F HFXMCM-rD2-R | 5'-GTCCAGGAGTCCCCC-GAGACGCCGCAGAGC-3' 5'-GCTCTGCGGCGTCTC-GGGGGACTCCTGGAC-3' |
| HFXMCM-rD3-F HFXMCM-rD3-R | 5'-CGCGTCCAGGAGTCC-ACGCCGCAGAGCATC-3' 5'-GATGCTCTGCGGCGT-GGACTCCTGGACGCG-3' |
| HFXMCM-rD4-F HFXMCM-rD4-R | 5'-CTGCGCGTCCAGGAG-CCGCAGAGCATCGAC-3' 5'-GTCGATGCTCTGCGG-CTCCTGGACGCGCAG-3' |
| C. FOR CONSTRUCTION OF β9-β10 β-HAIRPIN MUTANTS BY OEM | |
| HFXMCM-bH1-F HFXMCM-bH1-R | 5'-GTCGGCGTCCTCGCAGCGGAACAGCAGACATCG-3' 5'-CGATGTCTGCTGTTCCGCTGCGAGGACGCCGAC-3' |
| HFXMCM-bH2-F HFXMCM-bH2-R | 5'-GTCCTCCACATCGCAGCGCAGACATCGGGCAAC-3' 5'-GTTGCCCGATGTCTGCGCTGCGATGTGGAGGAC-3' |
| HFXMCM-bH3-F HFXMCM-bH3-R | 5'-CACATCGAACAGGCAGCGTCGGGCAACGAGAAG-3' 5'-CTTCTCGTTGCCCGACGCTGCCTGTTCGATGTG-3' |
| HFXMCM-bH4-F HFXMCM-bH4-R | 5'-GAACAGCAGACAGCAGCGAACGAGAAGACGCCC-3' 5'-GGGCGTCTTCTCGTTCGCTGCTGTCTGCTGTTC-3' |
| HFXMCM-bH5-F HFXMCM-bH5-R | 5'-CAGACATCGGGCGCAGCGAAGACGCCCGTCTTC-3' 5'-GAAGACGGGCGTCTTCGCTGCGCCCGATGTCTG-3' |
| HFXMCM-bH6-F HFXMCM-bH6-R | 5'-TCGGGCAACGAGGCAGCGCCCGTCTTCGACTAC-3' 5'-GTAGTCGAAGACGGGCGCTGCCTCGTTGCCCGA-3' |
| HFXMCM-hD1-F HFXMCM-hD1-R | 5'-CACATCGAACAGCAG-GGCAACGAGAAGACG-3' 5'-CGTCTTCTCGTTGCC-CTGCTGTTCGATGTG-3' |
| HFXMCM-hD2-F HFXMCM-hD2-R | 5'-CTCCACATCGAACAG-AACGAGAAGACGCCC-3' 5'-GGGCGTCTTCTCGTT-CTGTTCGATGTGGAG-3' |
| HFXMCM-hD3-F HFXMCM-hD3-R | 5'-GTCCTCCACATCGAA-GAGAAGACGCCCGTC-3' 5'-GACGGGCGTCTTCTC-TTCGATGTGGAGGAC-3' |
| HFXMCM-hD4-F HFXMCM-hD4-R | 5'-GGCGTCCTCCACATC-AAGACGCCCGTCTTC-3' 5'-GAAGACGGGCGTCTT-GATGTGGAGGACGCC-3' |
| D. FOR CONSTRUCTION OF ZINC BINDING DOMAIN MUTANTS BY OEM | |
| HFXMCM-C137A-F HFXMCM-C137A-R | 5'-CCGCCTTCGAGGCGCAGCGCTGCGG-3' 5'-CCGCAGCGCTGCGCCTCGAAGGCGG-3' |
| HFXMCM-C140A-F HFXMCM-C140A-R | 5'-AGTGCCAGCGCGCGGGGACGATGAG-3' 5'-CTCATCGTCCCCGCGCGCTGGCACT-3' |
| HFXMCM-C159A-F HFXMCM-C159A-R | 5'-AACCCCACGAGGCGCAGGGATGCGA-3'5'-TCGCATCCCTGCGCCTCGTGGGGTT-3' |
| HFXMCM-C162A-F HFXMCM-C162A-R | 5'-AGTGTCAGGGAGCGGAGCGCCAGGG-3' 5'-CCCTGGCGCTCCGCTCCCTGACACT-3' |
| E. FOR CONSTRUCTION OF SILENT Acc65I mutation by OEM | |
| HFXMCM-S1-F HFXMCM-S1-R | 5'-TGCCAGCGCTGCGGTACCATGAGCTACATC-3' 5'-GATGTAGCTCATGGTACCGCAGCGCTGGCA-3' |
| F. FOR DETECTION OF β7-β8 β-HAIRPIN MUTANTS | |
| HFXMCM-rM1-WT | 5'-CAGAAACTGCGCGTCCAG-3' |
| HFXMCM-rM1-MUT | 5'-CAGAAACTGCGCGTCGCC-3' |
| HFXMCM-rM2-WT | 5'-AAACTGCGCGTCCAGGAG-3' |
| HFXMCM-rM2-MUT | 5'-AAACTGCGCGTCCAGGCC-3' |
| HFXMCM-rM3-WT | 5'-GTCCAGGAGTCCCCCGAG-3' |
| HFXMCM-rM3-MUT | 5'-GTCCAGGAGTCCCCCGCC-3' |
| HFXMCM-rM4-WT | 5'-TCCCCCGAGGGCCTGCGC-3' |
| HFXMCM-rM4-MUT | 5'-TCCCCCGAGGGCCTGGCG-3' |
| HFXMCM-rM5-WT | 5'-GGCCTGCGCGGGGGCGAG-3' |
| HFXMCM-rM5-MUT | 5'-GGCCTGCGCGGGGGCGCC-3' |
| HFXMCM-rM6-WT | 5'-GGGGGCGAGACGCCGCAG-3' |
| HFXMCM-rM6-MUT | 5'-GGGGGCGAGACGCCGGCC-3' |
| HFXMCM-rD1-MUT | 5'-GGAGTCCCCCGAGGGCGAG-3' |
| HFXMCM-rD2-MUT | 5'-CCAGGAGTCCCCCGAGAC-3' |
| HFXMCM-rD3-MUT | 5'-CGTCCAGGAGTCCACGCC-3' |
| HFXMCM-rD4-MUT | 5'-CTGCGCGTCCAGGAGCCG-3' |
| G. FOR DETECTION OF β9-β10 β-HAIRPIN MUTANTS | |
| HFXMCM-bH1-WT | 5'-GTCGGCGTCCTCCACATC-3' |
| HFXMCM-bH1-MUT | 5'-GTCGGCGTCCTCGCAGCG-3' |
| HFXMCM-bH2-WT | 5'-GTCCTCCACATCGAACAG-3' |
| HFXMCM-bH2-MUT | 5'-GTCCTCCACATCGCAGCG-3' |
| HFXMCM-bH3-WT | 5'-CACATCGAACAGCAGACA-3' |
| HFXMCM-bH3-MUT | 5'-CACATCGAACAGGCAGCG-3' |
| HFXMCM-bH4-WT | 5'-GAACAGCAGACATCGGGC-3' |
| HFXMCM-bH4-MUT | 5'-GAACAGCAGACAGCAGCG-3' |
| HFXMCM-bH5-WT | 5'-CAGACATCGGGCAACGAG-3' |
| HFXMCM-bH5-MUT | 5'-CAGACATCGGGCGCAGCG-3' |
| HFXMCM-bH6-WT | 5'-ACATCGGGCAACGAGAAGA-3' |
| HFXMCM-bH6-MUT | 5'-ACATCGGGCAACGAGGCAG-3' |
| HFXMCM-hD1-WT | 5'-CACATCGAACAGCAGACA-3' |
| HFXMCM-hD1-MUT | 5'-CACATCGAACAGCAGGGC-3' |
| HFXMCM-hD2-WT | 5'-CTCCACATCGAACAGCAG-3' |
| HFXMCM-hD2-MUT | 5'-CTCCACATCGAACAGAAC-3' |
| HFXMCM-hD3-WT | 5'-TCCTCCACATCGAACAGC-3' |
| HFXMCM-hD3-MUT | 5'-TCCTCCACATCGAAGAGA-3' |
| HFXMCM-hD4-WT | 5'-GGCGTCCTCCACATCGAA-3' |
| HFXMCM-hD4-MUT | 5'-GGCGTCCTCCACATCAAG-3' |
| H. FOR DETECTION OF ZINC BINDING DOMAIN MUTANTS | |
| HFXMCM-C137A-WT | 5'-GAAGCCGCCTTCGAGTGC-3' |
| HFXMCM-C137A-MUT | 5'-GAAGCCGCCTTCGAGGCG-3' |
| HFXMCM-C140A-WT | 5'-TTCGAGTGCCAGCGCTGC-3' |
| HFXMCM-C140A-MUT | 5'-TTCGAGTGCCAGCGCGCG-3' |
| HFXMCM-C159A-WT | 5'-CAGGAACCCCACGAGTGT-3' |
| HFXMCM-C159A-MUT | 5'-CAGGAACCCCACGAGGCG-3' |
| HFXMCM-C162A-WT | 5'-CACGAGTGTCAGGGATGC-3' |
| HFXMCM-C162A-MUT | 5'-CACGAGTGTCAGGGAGCG-3' |
| I. REVERSE PRIMER FOR MUTANT DETECTION | |
| HFXMCM-R1150 | 5'-GCGAATCCGCGAGCCGTC-3' |
Restriction sites in oligonucleotide primers used for pTA131-HfxMCM-HXba construction underlined. Oligonucleotide primers for overlap extension mutagenesis (OEM) are shown in top strand-bottom strand pairs with the mutated bases underlined in the top strand primer (for amino acid substitutions) or with deletion boundaries indicated with a dash. Oligonucleotide primers for mutant detection are shown with 3 ' mismatched bases underlined.