TABLE 3.
PCR primers used in this study
| Primer | Sequence (5′–3′)a | Purpose | Reference |
|---|---|---|---|
| BACTH construct primers | |||
| M1_36_pUT18_F | catgcatgAAGCTTgCGTAACTCACTGGGTCGAAA | Used to construct mcpMΔ36 | This study |
| M1_36_pUT18_R | catgcatgGAATTCggTCGGTTACATGTTCCGC | ||
| M1_36_pKNT25_F | catgcatgAAGCTTgCGTAACTCACTGGGTCGAAA | Used to construct mcpMΔ36 | This study |
| M1_36_pKNT25_R | catgcatgGAATTCggTCGGTTACATGTTCCGC | ||
| M1_36_pUT18C_F | catgcatgTCTAGAgCGTAACTCACTGGGTCGAAA | Used to construct mcpMΔ36 | This study |
| M1_36_pUT18C_R | catgcatgGAATTCTTATCGGTTACATGTTCCGC | ||
| M1_36_pKT25_F | catgcatgTCTAGAgCGTAACTCACTGGGTCGAAA | Used to construct mcpMΔ36 | This study |
| M1_36_pKT25_R | catgcatgGAATTCTTATCGGTTACATGTTCCGC | ||
| mcpI_pUT18_F | catgcatgAAGCTTgATGGAGGGCGCTACTATGTT | Used to construct mcpI | This study |
| mcpI_pUT18_R | catgcatgGAATTCggTTTCGCGGAGATTGTTCTT | ||
| mcpI_pKNT25_F | catgcatgAAGCTTgATGGAGGGCGCTACTATGTT | Used to construct mcpI | This study |
| mcpI_pKNT25_R | catgcatgGAATTCggTTTCGCGGAGATTGTTCTT | ||
| mcpI_pUT18C_F | catgcatgTCTAGAgATGGAGGGCGCTACTATGTT | Used to construct mcpI | This study |
| mcpI_pUT18C_R | catgcatgGAATTCCTATTTCGCGGAGATTGTTCTT | ||
| mcpI_pKT25_F | catgcatgTCTAGAgATGGAGGGCGCTACTATGTT | Used to construct mcpI | This study |
| mcpI_pKT25_R | catgcatgGAATTCCTATTTCGCGGAGATTGTTCTT | ||
| ompF_pUT18_F | catgcatgAAGCTTgATGATGAAGCGCAATATTCTGG | Used to construct ompF | This study |
| ompF_pUT18_R | catgcatgGAATTCggGAACTGGTAAACGATACCC | ||
| ompF_pKNT25_F | catgcatgAAGCTTgATGATGAAGCGCAATATTCTGG | Used to construct ompF | This study |
| ompF_pKNT25_R | catgcatgGAATTCggGAACTGGTAAACGATACCC | ||
| ompF_pUT18C_F | catgcatgTCTAGAgATGATGAAGCGCAATATTCTGG | Used to construct ompF | This study |
| ompF_pUT18C_R | catgcatgGAATTCTTAGAACTGGTAAACGATACCC | ||
| ompF_pKT25_F | catgcatgTCTAGAgATGATGAAGCGCAATATTCTGG | Used to construct ompF | This study |
| ompF_pKT25_R | catgcatgGAATTCTTAGAACTGGTAAACGATACCC | ||
| M13R49 | GAGCGGATAACAATTTCACACAGG | Used to verify BACTH constructs | This study |
| pUT18C_R | GCATCAGAGCAGATTGTACTGAGAG | ||
| pUT18_R | TTCGCGATCCAGGCCGC | ||
| pKT25_R | CCAGGGTTTTCCCAGTCACG | ||
| pKNT25_R | GCGATTGCTGCATGGTCA | ||
| mcpI construct primers | |||
| mcpI_flag_F | ccgGAATTCATGGAGGGCGCTACTAT | Used to construct pMMB207::mcpI with addition of a Flag tag in the C terminus | This study |
| mcpI_flag_R | acgcGTCGACCTACTTGTCGTCATCGTCTTTGTAGTCTTTCGCGGAGATTGTTCT | ||
| Gene deletion mutant primers | |||
| A1_mcpMI_KO_F | ggaAGATCTATGGCAATGCTGGAAGAG | Used to construct suicide plasmid pDM4ΔmcpM/ΔmcpI(A1+A2) to delete mcpM and mcpI | This study |
| A1_mcpMI_KO_R | AATCTTGCCGATCATATTTCCCCTATCGGT | ||
| A2_mcpMI_KO_F | GAAATATGATCGGCAAGATTCATGGACTA | ||
| A2_mcpMI_KO_R | acgcGTCGACCTTCATAATACGGAACTGTCAG | ||
| mcpM_F | AGATGAGATAACGCTTGTCA | Used to confirm mcpM deletion and complementation | 7 |
| mcpM_R | ACTTCCTCTGTTACCACTTC | ||
| mcpI_F | TATGTGGTTTGTTACTGGGAT | Used to confirm mcpI deletion and complementation | 7 |
| mcpI_R | CGCGGAGATTGTTCTTATTT | ||
| SacI_pMIADB | ggtggtGAGCTCTGGAACGAGATGTACTGAACAGAGGCCGTG | Used to construct pmic-210/0mcpMIADB | This study |
| HindIII_MIADB | ggtggtAAGCTTTCACCTCCTGTTGGGGTGATTATTTATATTATGAGTTCTGGC | ||
| Construct tdTomato primers | |||
| tdtomato_Xbal | tgcTCTAGATTTAAGAAGGAGATATACATATGGTGAGCAAGGGCGAGGA | Construct pFPV-tdTomato for labeling bacteria | This study |
| tdtomato_SphI | catGCATGCCTACTTGTACAGCTCGTCCATGC | ||
| Bacteriocin primers | |||
| mcmM_F | GCATTAGTTGGGGAGCCAGA | Used to detect the gene mcmM (microcin M) | This study |
| mcmM_R | CAACCCCACCAGGAACAGTT | ||
| mchB_F | TGCGAGAAATAACAGAATCACAGT | Used to detect the gene mchB (microcin H47) | This study |
| mchB_R | CCAGCAGAAGAACTGGCACT | ||
| mceA_F | CTTGGCCCGATGAGTACAGG | Used to detect the gene mceA (microcin E492) | This study |
| mceA_R | TTTTGGTGCAGGAGAGACCG | ||
| cvaC_F | TTCAGGGCGTGATATTGCGA | Used to detect the gene cvaC (microcin V) | This study |
| cvaC_R | TCCCGCAGCATAGTTCCATG | ||
| cia_F | TGTAAACCCTCCACGTGTCG | Used to detect the gene cia (colicin 1a) | This study |
| cia_R | GACAACCGGGTGTCCAGAAT | ||
| cib_F | GGTGGTGACAGAGGATGTGG | Used to detect the gene cib (colicin 1b) | This study |
| cib_R | GTCTCCGCCATATGGACCTG | ||
| mcbA_F | TGGAATTAAAAGCGAGTGAATTT | Used to detect the gene mcbA (microcin B17) | This study |
| mcbA_R | CACCGTTTCCACCACTACAA | ||
| ompF sequence primers | |||
| ompF_up | CGCTATCAGGGTAACGGGAG | Used to sequence ompF in E. coli | This study |
| ompF_down | AGCACTTTCACGGTAGCGAA | ||
a
Restriction enzyme sites are underlined.