TABLE 2.
Oligonucleotides used in this study
| Primer name | Sequence (5′ to 3′)a | Restriction site | Purpose |
|---|---|---|---|
| FPAsefw | GGATTATGTCAAAAGGAGAAGAATTATTTACAG | AseI | Amplification of fluorescent |
| FPBamrv | GGATCCTTACTTATATAATTCATCCATTCCGTG | BamHI | protein genes |
| GFPAsefw | GGATTAATGAGTAAAGGAGAAGAACTTTTCACTG | AseI | Amplification of non-codon- |
| GFPBamrv | GGATCCTTATTTGTATAGTTCATCCATGCCA | BamHI | optimized gfpmut1 |
| CFPAsefw | GGATTAATGGTGAGCAAGGGCGA | AseI | Amplification of non-codon- |
| CFPBamrv | GGATCCTTACTTGTACAGCTCGTCCATG | BamHI | optimized cfp |
| YFPAsefw | GGATTAATGGTGAGCAAGGGCGAG | AseI | Amplification of non-codon- |
| YFPBamrv | GGATCCTTACTTGTACAGCTCGTCCATGC | BamHI | optimized yfp |
| SDMCFPV2 | TATACAAAAAGGAGAACGCATAATGGTTTCAAAAGGAGAAGAATTATTTACAG | None | Site-directed mutagenesis of cfpopt to introduce valine-2 (CFPopt) |
| SDMCFPH-L | TTCGTAACAGCAGCAGGAATTACACTAGGAATGGATGAATTATATAAGTAAG | None | Site-directed mutagenesis of gfpopt for exchange H232L (CFPopt) |
| SDMCFPSY-TW | GGCCCACACTTGTGACTACTTTAACATGGGGAGTACAATGTTTTTC | None | Site-directed mutagenesis of gfpopt for exchanges S65T and Y66W (CFPopt) |
| SDMCFPN-I | GGACATAAATTAGAATACAATTACATCAGTCATAACGTATACATAATGGCA | None | Site-directed mutagenesis of gfpopt for exchange N146I (CFPopt) |
| SDMCFPM-T | AGAATACAATTACAACAGTCATAACGTATACATAACGGCAGATAAACAGAAAAA | None | Site-directed mutagenesis of gpfopt for exchange M153T (CFPopt) |
| SDMYFPSV-GL | ATGGCCCACACTTGTGACTACTTTAGGATACGGACTACAATGTTTTTCAAGATATC | None | Site-directed mutagenesis of gfpopt for exchanges S65G and V68L (YFPopt) |
| SDMYFPS-A | TTTAACATACGGAGTACAATGTTTTGCAAGATATCCAGATCATATGAAAC | None | Site-directed mutagenesis of gfpopt for exchange S72A (YFPopt) |
| SDMYFPT-Y | CTTCTACCAGATAATCATTACTTAAGCTATCAAAGCGCTTTATCTAAAGACCCAAA | None | Site-directed mutagenesis of gfpopt for exchange T203Y (YFPopt) |
| SDMPAGFPLT-FS | GGCCCACACTTGTGACTACTTTTTCATACGGAGTACAATGTTTTTC | None | Site-directed mutagenesis of gfpopt for exchanges L64F and T65S (PAGFPopt) |
| SDMPAGFPS-A | GGCAGATAAACAGAAAAATGGAATTAAAGCTAACTTTAAAATAAGACACAATATAGAAG | None | Site-directed mutagenesis of gfpopt for exchange V163A (PAGFPopt) |
| SDMPAGFPT-H | CTTCTACCAGATAATCATTACTTAAGCCATCAAAGCGCTTTATCTAAAGACCCAAA | None | Site-directed mutagenesis of gfpopt for exchange T203H (PAGFPopt) |
| pSW4Seq | AAAGTTCTGTTTAAAAAGCCAAAAAT | None | Sequencing oligonucleotide for verification of pSW4 inserts |
| M13f | GTAAAACGACGGCCAGT | None | Sequencing oligonucleotide for verification of pCR2.1 inserts |
| M13r | AACAGCTATGACCATG | None | Sequencing oligonucleotide for verification of pCR2.1 inserts |
| S10fwAse | GGATTAATATTCACCACCGTTCTTATGACTA | AseI | Forward primer to amplify S10 promoter from S. aureus |
| S10CFPrv | CTTGCTCACCATAATTTTCCCTCCTTATTCGTCTA | None | Reverse primer for S10 promoter with cfp overlap |
| S10CFPoptrv | CTTTTGAAACCATAATTTTCCCTCCTTATTCGTCTA | None | Reverse primer for S10 promoter with cfpopt overlap |
| CFPoptBclfw | ATTGATCATACAAAAAGGAGAACGCATAATGGTGAGCAAGGGCGA | BclI | Amplification of cfpopt with |
| CFPoptBamrv | GGATCCTTACTTATATAATTCATCCATTCCTAGTG | BamHI | ribosomal binding site for pTetON cloning |
| YFPoptBclfw | ATTGATCATACAAAAAGGAGAACGCATAATGTCAAAAGGAGAAGAATTATTTACAG | BclI | Amplification of yfpopt with ribosomal binding site for |
| YFPoptBamrv | CGGGATCCTTACTTATATAATTCATCCATTCCGTG | BamHI | pTetON cloning |
| CFPSOEfw | GGAGGGAAAATTATGGTGAGCAAGGGCGA | None | Forward primer for cfp with S10 promoter overlap |
| CFPoptSOEfw | GGAGGGAAAATTATGGTTTCAAAAGGAGAAGAATT | None | Forward primer for cfpopt with S10 promoter overlap |
| GyrAfw | AAAACCTGTGCATCGTAGGG | None | Forward primer for gyrA |
| GyrArv | ACATTAGCATTGGCATCACG | None | Reverse primer for gyrA |
a
Restriction sites are in boldface, and overlaps are in italic.