The pGRT1 plasmid of Pseudomonas putida DOT-T1E encodes functions relevant for survival under harsh conditions in the environment
Article first published online: 23 MAY 2011
© 2011 Society for Applied Microbiology and Blackwell Publishing Ltd
Thematic Issue: Extremophiles. Guest Editors: Ricardo Cavicchioli, Ricardo Amils, Dirk Wagner, Terry McGenity
Volume 13, Issue 8, pages 2315–2327, August 2011
How to Cite
Molina, L., Duque, E., Gómez, M. J., Krell, T., Lacal, J., García-Puente, A., García, V., Matilla, M. A., Ramos, J.-L. and Segura, A. (2011), The pGRT1 plasmid of Pseudomonas putida DOT-T1E encodes functions relevant for survival under harsh conditions in the environment. Environmental Microbiology, 13: 2315–2327. doi: 10.1111/j.1462-2920.2011.02492.x
- Issue published online: 21 AUG 2011
- Article first published online: 23 MAY 2011
- Received 16 March, 2011; accepted 22 March, 2011.
Fig. S1. A. Southern blot of the total DNA from the wild-type and mutant ORF49:: ΩKm. DNA was digested with BamHI and the blot was hybridized with a DIG-labelled probe synthesized by amplification using Rep1 and Rep2 primers. B. Southern blot analysis of the total DNA digested with ScaI from wild-type and mutants ORF72: ΩKm (mcpT-1) and ORF97: ΩKm (mcpT-2). The blot was hybridized with a Dig-labelled mcp probed synthesized by PCR with the Mcp-1 and Mcp-2 oligonucleotides.
Fig. S2. Putative RNA regulatory element of the antisense-RNA regulated loci. A. Basic RNA unit that acts as antitoxin in Erwinia carotovora. B. Upstream DNA of the toxN gene are the basic RNA units; the first ATG of the ToxN protein is highlighted by a circle.
Fig. S3. Survival of P. putida DOT-T1E, P. putida DOT-T1E-100 and DOT-T1E-ORF34:: ΩKm after exposure to ultraviolet light. Serial dilutions of the cultures of the different strains were drop-plated and allowed to dry. Then plates were exposed to ultraviolet light (254 nm) for 10 s, and incubated overnight at 30°C.
Fig. S4. A. CAS assay. Siderophore production was determined by the production of a color change in CAS plates from blue to orange P. aeruginosa PAO1 and Bacillus subtillis were used as positive and negative controls. The four P. putida strains submitted to the assay were DOT-T1E, DOT-TIE100 – deficient in pGRT1; a pGRT1 mutant with a knock-out at ORF35 and this mutant complemented with the orf35 gene cloned in pBBRMCS5 plasmid. B. Absorption spectra of the supernatants of cultures of the wild-type strain grown in the absence and in the presence of iron. C) Absorption spectra of supernatants of the different cultures grown in absence of iron.
Table S1. Set of genes interrupting the plasmid backbone. Listed are genes found between coordinates 18927 and 41331 together with their relevant homologies. Other details are shown in Fig. 1.
Table S2. Genes borne on the Tn4653-like transposon. Genes highlighted in dark grey are within the two 7.6 inverted repeat sequences. Light grey indicates genes within the insertion sequence ISPpu12.
Table S3. Relevant genes involved in replication and maintenance of plasmid pGRT1. Genes that are homologs to the TA system are highlighted in grey.
Table S4. Oligonucleotides used to amplify DNA for construction of mutants. The sequences are given in the 5'-> 3' direction and the size of the amplicon is indicated in base pairs (bp).
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