Article Figures & Data
Figures
- FIG 1
Functional categories of the 398 genes identified as being required for growth in rich medium.
- FIG 2
Conditional lethal phenotype of the rhamnose-inducible BCAM0746 mutant. Undiluted cultures of mutants grew visibly on plates supplemented with either glucose or rhamnose prior to depletion of the existing protein; however, at 100- and 1,000-fold dilutions, the mutant grew well in the presence of rhamnose (a) but was unable to grow on plates supplemented with glucose (b). Addition of 1 mM l-arginine partially restored growth of the BCAM0746 conditional mutant in the presence of glucose (c), and addition of 10 mM l-arginine restored growth to the level of the induced mutant (d). The J2315 BCAM0746 conditional mutant grew equally well in the presence of rhamnose (e) and glucose (f).
- FIG 3
Conditional lethal phenotype of rhamnose-inducible mutants of BCAM0632, BCAM0986, and BCAM0998 on ABC minimal medium. The BCAM0632 conditional mutant had levels of growth comparable to those of the B. cenocepacia H111 wild type. The BCAM0986 conditional mutant was unable to grow on ABC medium, and the BCAM0998 conditional mutant had reduced growth at the 10−2 and 10−3 dilutions compared with the wild-type strain H111.
- FIG 4
Visualization of the numbers and positions of transposon insertions within the coding sequence and the upstream region of RNase E (I35_RS04905). Note that no insertions were found within a large part of the upstream intergenic region of this gene and that the first insertions are found only approximately 1,200 bp downstream of the annotated start of the reading frame. This gene is duplicated in B. cenocepacia J2315, with each functional copy being designated BCAL0982 and BCAL2888. Image taken from the LB data set.
Tables
- TABLE 1
Numbers of unique insertions, average insertion frequencies, and numbers of essential genes of the three B. cenocepacia H111 replicons in rich medium
Chromosome Size of coding regions (bp) No. of unique insertions Insertion frequency (bp) No. of essential genes 1 3,099,313 496,269 6 358 2 2,657,050 288,110 9 30 3 904,321 72,122 12 10 - TABLE 2
Validation of H111 candidate essential genesa
Conditional mutant J2315 identifier H111 RefSeq identifier Function Essentialb B. cenocepacia BCAM0632 I35_RS19210 N-Acetyltransferase N BCAM0746-strain H111 I35_RS19780 Arginosuccinate synthase Y BCAM0746-strain J2315 I35_RS19780 Arginosuccinate synthase N BCAM0911 I35_RS20630 1-Deoxy-d-xylulose-5-phosphate synthase Y BCAM0913 I35_RS20640 tRNA threonylcarbamoyladenosine modification protein TsaD Y BCAM0918 I35_RS20665 RNA polymerase sigma factor RpoD Y BCAM0967 I35_RS20915 Succinate dehydrogenase Y BCAM0972 I35_RS20940 Type II citrate synthase Y BCAM0986 I35_RS21020 Aspartate-semialdehyde dehydrogenase N BCAM0994 I35_RS21060 Acetyl-CoAc carboxylase carboxyltransferase subunit beta Y BCAM0995 I35_RS21065 Bifunctional folylpolyglutamate synthase/dihydrofolate synthase Y BCAM0998 I35_RS21080 Amidophosphoribosyltransferase N BCAM1362 I35_RS22520 Cell division protein Y BCAM1931 I35_RS25190 Porin N BCAM2650 I35_RS29140 trans-2-Enoyl-CoA reductase Y BCAM2688 I35_RS29330 N-Acylglucosamine 2-epimerase N B. thailandensis E264/J2315 orthologue BTH_I0484/BCAM0545 I35_RS18760 PTS fructose transporter subunit IIA N BTH_I1642/BCAM0855 I35_RS20355 UDP-glucose 6-dehydrogenase N BTH_II0656/BCAM0963 I35_RS20895 Hypothetical protein N BTH_II1941/BCAM1812 I35_RS24870 Agmatinase N BTH_II1863/BCAM2321 I35_RS27455 Electron transfer flavoprotein subunit alpha N BTH_II0780/BCAM2358A I35_RS27645 Hypothetical protein N ↵a The conditional rhamnose-inducible mutants were spotted on LB plates supplemented with either 0.5% rhamnose (permissive condition) or 0.5% glucose (nonpermissive condition) as described in Materials and Methods. Growth was recorded after 24 h of incubation at 37°C.
↵b Y, essential; N, nonessential.
↵c CoA, coenzyme A.
- TABLE 3
Comparison of the numbers of transposon insertions found in the flagellar biosynthesis genes in rich medium, minimal medium and microoxiaa
Gene Function No. of unique insertions LB ABC Microoxia flgL Flagellar hook-associated protein FlgL 303 184 21 flgK Flagellar hook protein FlgK 522 325 47 —b Flagellar brake protein YcgR 158 77 776 flgJ Flagellar rod assembly protein FlgJ 213 130 27 — Flagellar P-ring protein FlgI 281 153 29 — Flagellar L-ring protein 49 24 3 flgG Flagellar basal body rod protein FlgG 231 108 25 flgF Flagellar basal body rod protein FlgF 174 57 10 flgE Flagellar hook protein FlgE 328 160 18 — Flagellar basal body rod modification protein FlgD 106 56 8 flgC Flagellar basal body rod protein FlgC 106 60 2 flgB Flagellar biosynthesis protein FlgB 91 40 3 — Flagellar basal body P-ring biosynthesis protein FlgA 164 58 6 — Flagellar biosynthesis protein FlgM 87 69 31 — Flagellar synthesis chaperone protein FlgN 113 58 52
Supplemental material
- Supplemental file 1 -
Table S1 (Details of 398 genes identified as essential in LB rich medium)
XLSX, 658K
- Supplemental file 2 -
Table S2 (Comparison of B. cenocepacia J2315 and H111 essential genes)
XLSX, 37K
- Supplemental file 3 -
Table S3 (Data on essential genes from Bloodworth et al. and the LB rich medium Tn-Seq data set)
XLSX, 14K
- Supplemental file 4 -
Table S4 (Genes identified as essential in ABC medium)
XLSX, 632K
- Supplemental file 5 -
Table S5 (Comparison of genes unable to grow in ABC medium with essential genes identified by Tn-Seq)
XLSX, 11K
- Supplemental file 6 -
Table S6 (Genes identified as essential in 0.5% oxygen conditions)
XLSX, 662K
- Supplemental file 7 -
Table S7 (Details of core essential genome of B. cenocepacia H111)
XLSX, 53K
- Supplemental file 8 -
Tables S8 (Bacterial strains and plasmids used) and S9 (Oligonucleotides used)
PDF, 299K
- Supplemental file 9 -
Fig. S10 (UID values and frequency)
PDF, 63K
- Supplemental file 10 -
Captions to Tables S1 to S9 and Fig. S10
PDF, 294K
- Supplemental file 1 -
