TABLE 2.
Primers used in this study
| Primer | Sequencea (5′–3′) | Target gene | REb |
|---|---|---|---|
| For cat transcriptional fusion | |||
| phoH-Fw41 | CGAGGATCCTGGATATGGGGTTGCTGTTTG | phoHEc | BamHI |
| phoH-Rv42 | CGAAAGCTTCATGGAGAGCACCTTGAGTTG | phoHEc | HindIII |
| phoH-Fw | CGAGGATCCAATATGGCTGGCTGGATCTG | phoHS Tm | BamHI |
| phoH-Rv14 | CGAAAGCTTGTCAGTCTCTTACAGAAAGATTAC | phoHS Tm | HindIII |
| phoH-Fw15 | CGAGGATCCTGTAAGAGACTGACAATGACGC | phoHS Tm | BamHI |
| phoH-Rv | CGAAAGCTTCCATGGATAGCACCTTGAGT | phoHS Tm | HindIII |
| phoH-Fw27 | CGAGGATCCTGACGCAATAGAGTAATGACAAAA | phoHS Tm | BamHI |
| phoH-Fw37 | GATCCCCCGCAGTAGCTAATGATTATCTTTTTTAGTCTCCTGCCGATGAAATAATCGTGTAATCTTTCTGTAAGAA | phoHS Tm | |
| phoH-Rv38 | AGCTTTCTTACAGAAAGATTACACGATTATTTCATCGGCAGGAGACTAAAAAAGATAATCATTAGCTACTGCGGGG | phoHS Tm | |
| phoH-Fw79 | CAGCATTAATACCCGCAGTAGAAAGCACACAGCTTTTTTAGTCTCCTGCCGATG | phoHS Tm | |
| phoH-Rv80 | CATCGGCAGGAGACTAAAAAAGCTGTGTGCTTTCTACTGCGGGTATTAATGCTG | phoHS Tm | |
| phoH-Fw83 | CTGCCGATGAAATAATCGTGCGGCTCTTCTGTAAGAGACTGACAATG | phoHS Tm | |
| phoH-Rv84 | CATTGTCAGTCTCTTACAGAAGAGCCGCACGATTATTTCATCGGCAG | phoHS Tm | |
| phoH-Fw17 | CGAGGATCCAGCGATGGGAGAGAGGACAC | phoHS Tm | BamHI |
| pstS-Fw71 | CGAGGATCCCTCTTTGTCCTGGCGATCCC | pstSS Tm | BamHI |
| pstS-Rv72 | CGAAAGCTTGAACAGGCCTTCCTGATTCAG | pstSS Tm | HindIII |
| For gene deletion | |||
| phoH-H1P1 | GGCTTACAAGGAAGCCAACCCTCAGATGTGTGTGCGCATAATTGTAGGCTGGAGCTGCTTCG | phoHS Tm | |
| phoH-H2P2 | CGGGCTCTGAAAGTCAATGCTATATTAGCTGTATGCGTGAAGCATATGAATATCCTCCTTAG | phoHS Tm | |
| siiE-H1P1 | ATGGGAAATAAAAGCATACAAAAGTTTTTTGCCGATCAAAATTGTAGGCTGGAGCTGCTTCG | siiE | |
| siiE-H2P2 | TTATGCGTGTTCTTCTTGATTATCTACAGGTAGCGTAACTTCCATATGAATATCCTCCTTAG | siiE | |
| SphoB H1P1 | CGACTAACAGGGCAAATTATGGCGAGACGTATTCTGGTCGTATGTAGGCTGGAGCTGCTTCG | phoBS Tm | |
| SphoB H2P2 | CAACTGGCGGAAAAGGCATTAAAAGCGGGTCGAAAAACGATACATATGAATATCCTCCTTAG | phoBS Tm | |
| For gene FLAG tagging | |||
| phoHSFlagFw | GCGCTCGGCGCTTTGTCAGCGAACGCTTCACGCATACAGCGACTACAAAGACCATGACGGT | phoHS Tm | |
| phoHSFlagRv | ACAAAGCCCGGTTTCCCGGGCTCTGAAAGTCAATGCTATACATATGAATATCCTCCTTAGTTC | phoHS Tm | |
| phoHEcFlagFw | CGTTCGGCACTTTGCCAACGTACGCTGCATGCCTACAGTGACTACAAAGACCATGACGGT | phoHEc | |
| phoHEcFlagRv | ACAAAGCCCGGTTCGCCCGGGCTCTGCACCGATAACACACCATATGAATATCCTCCTTAGTTC | phoHEc |
a
Underlined letters indicate the respective restriction enzyme site in the primer. The sequences corresponding to the template pKD4 or pSUB11 plasmids (Table 1) are in italics.
b
RE, restriction enzyme for which a site was generated in the primer.