TABLE 1.
Strains, plasmids, and oligonucleotides used in this study
| Strain, plasmid, or primer | Description or sequencea | Reference, source, or purpose |
|---|---|---|
| Strains | ||
| Caulobacter crescentus | ||
| NA1000 | Synchronizable derivative of CB15 | 53 |
| NA1000 FLAG-RNase E | NA1000 expressing FLAG-RNase E | 55 |
| MM50 | NA1000 ΔrhlB | This work |
| MM71 | NA1000 FLAG-RNase E ΔrhlB | This work |
| MM74 | NA1000 rhlE::mini-Tn5 | 29 |
| MM82 | NA1000 ΔrhlB rhlE::mini-Tn5 | This work |
| MM84 | NA1000 FLAG-RhlE | This work |
| Escherichia coli | ||
| DH5α | (ϕ80lacZΔM15) hsdR17 recA1 endA1 gyrA96 thi-1 relA1 | 63 |
| S17-1 | 294::RP4-2(Tc::Mu)(Km::Tn7) | 56 |
| Plasmids | ||
| pGEM | Cloning vector, Apr | Promega |
| pNPTS138 | Suicide vector containing oriT and sacB, Kanr | M. R. K. Alley, unpublished data |
| pET-15 | Cloning vector | Novagen |
| pET-DUET | Cloning vector | Novagen |
| pBV-MCS4 | Cloning vector | 58 |
| pRKlacZ290 | Reporter vector for transcriptional fusions to lacZ, Tetr | 59 |
| pJBZ281 | Reporter vector for translational fusions to lacZ, Kanr | 60 |
| Primers | ||
| CC1847-C | TTGGGCCCCGTCGAAGGCGGGAGTCGTG | Deletion of rhlB |
| CC1847-D | TTTAAGCTTGGAGCCGCCCGTGCTTGG | Deletion of rhlB |
| CC1847-E | TTTAAGCTTTCGGCAAGATCCATCGAGG | Deletion of rhlB |
| CC1847-F | TTGAATTCCGGTGCGGTGTTCATGCCCT | Deletion of rhlB |
| RhlE1 | TGAATTCGACTGAAGATGACGGTTTCG | Amplification of rhlE promoter for transcriptional fusion |
| RhlE2 | TGGATCCGGTCGGTTCCTTGTAGC | Amplification of rhlE promoter for transcriptional fusion |
| P0835-1 | TGGATCCGGCGCGGGGTCCGTATCGG | Amplification of rhlE promoter without 5′ UTR for transcriptional fusion |
| RhlE 10 | CCATGGACTACAAGGACGACGACGACAAGGTGACTCAATTTTCCGACCTTGG | Construction of FLAG-RhlE strain |
| RhlE 11 | CTTGTCGTCGTCGTCCTTGTAGTCCATGGGTGTTGTGTCTTTCGTGCGCC | Construction of FLAG-RhlE strain |
| RhlE 12 | AAAAGAATTCGTGGCAGCTTGGCCATCTCCG | Construction of FLAG-RhlE strain |
| RhlE 13 | AGCTGCTTGTCGTTGCGACGG | Construction of FLAG-RhlE strain |
| RhlE-tra | AGGATCCACGTGTGTTGTGTCTTTCG | Amplification for rhlE translational fusion |
| ccRhlB-DUET.f | GCCATATGACTGAATTCACCGACC | Amplification of rhlB for cloning |
| ccHel.REV2 | GCCTCGAGCTACTTCGCGCCGCGCGGCGGCCGCGCGAGG | Amplification of rhlB for cloning |
| cc0835_Nde.F | CAGCCATATGACTCAATTTTCCGACCTTGGCCTG | Amplification of rhlE for cloning |
| cc0835_Bam.R | GCCGGATCCTTAGTCGATAGGCGACCAGCG | Amplification of rhlE for cloning |
a
Underlined letters indicate restriction enzyme recognition sites used for cloning purposes. Apr, ampicillin resistance; Kanr, kanamycin resistance; Tetr, tetracycline resistance.