TABLE 2.
Oligonucleotides and primers used in this study
| Oligonucleotide or primer | Sequencea | Relevant features |
|---|---|---|
| Oligonucleotides | ||
| ForSG38P | GATATAATccCGGgGCGTTTACAAAAACAG | Binds λ nt 45289–45418; mutates Gly 38 to Pro; creates a SmaI site |
| RevSG38P | GTAAACGCcCCGggATTATATCTGCCGCG | Reverse primer used in conjunction with ForSG38P |
| PETHis | ggaattccatatgagaaccatggtgatggtgatggtgcatgtaTATATCTCCTTCTTAAAGTTAAAC | Anneals downstream of NdeI site in pET11a and primes backward toward the T7 promoter; destroys the original NdeI site; used to create pETNH |
| RevT7 | cggaatTCATAAGTGCGGCGACG | Anneals to pET11a lacI flanking region; primes into lacI gene; used to create pETNH |
| Seamless Cloning primers | ||
| ForpKBEam | agttactcttcATAATCAACGTAAGGCGTTCC | Forward primer for generating vector fragments of transactivation plasmids |
| RevpKBEam | agttactcttcaGCATCTTCAGGTCTTACCCCC | Reverse primer for generating vector fragments of trans activation plasmids |
| ForS105Eam | agttactcttcaTATGCCAGAAAAACATGACCTG | Forward primer for generating S105 alleles with wild-type N termini |
| RevSEam | agttactcttcaATTATTGATTTCTACCATCTTCTAC | Reverse primer for generating S alleles with wild-type C termini |
| ForNHS105Eam | agttactcttcaCATGCACCATCACCATCACC | Forward primer for generating N-terminally tagged S105 alleles |
| RevCHSEam | agttactcttcaCTTAGTGATGGTGATGGTGATG | Reverse primer for generating C-terminally tagged S alleles |
| ForPhe94His | agttactcttctatcaccatcacggcggcGCTGCTAAAAAAGCCGGAG | Forward primer for generating an oligohistidine tag behind codon 94 |
| RevPhe94His | agttactcttctgatgatggtggccgccGAAGCGTTTGATAAGCGAACC | Reverse primer to be used in conjunction with ForPhe94His |
a
Lowercase letters represent either mismatches or insertions, and new restriction sites are indicated by bold type.