• Genome sequence of Ensifer adhaerens OV14 provides insights into its ability as a novel vector for the genetic transformation of plant genomes

      Rudder, Steven; Doohan, Fiona; Creevey, Christopher J.; Wendt, Toni; Mullins, Ewen; Science Foundation Ireland; 11/RFP.1/GEN/3420. (Biomed Central, 07/04/2014)
      Additional file 1:Figure S1. Circular representation of the four replicons of E. adhaerens OV14. Figure S2. Synteny plots showing total sequence of Ensifer adhaerens OV14 pOV14 (top bar) vs Agrobacterium tumefaciens C58 pTi (bottom bar), computed using DoubleACT version2 on tBLASTx setting with cut off set at 100. Visualised in Artemis ACT. Additional file 2; BLAST of Ensifer adhaerens OV14 replicons.xlsx. Excel file includes tables of BLAST search of individual Ensifer adhaerens OV14 replicons Additional file 3. BLAST of Agrobacterium tumefaciens C58 replicons.xlsx. Excel file includes tables of BLAST searches of individual Agrobacterium tumefaciens C58
    • Genome sequence of Ensifer adhaerens OV14 provides insights into its ability as a novel vector for the genetic transformation of plant genomes

      Rudder, Steven; Doohan, Fiona; Creevey, Christopher J.; Wendt, Toni; Mullins, Ewen (Biomed Central, 2014-04-07)
      Abstract Background Recently it has been shown that Ensifer adhaerens can be used as a plant transformation technology, transferring genes into several plant genomes when equipped with a Ti plasmid. For this study, we have sequenced the genome of Ensifer adhaerens OV14 (OV14) and compared it with those of Agrobacterium tumefaciens C58 (C58) and Sinorhizobium meliloti 1021 (1021); the latter of which has also demonstrated a capacity to genetically transform crop genomes, albeit at significantly reduced frequencies. Results The 7.7 Mb OV14 genome comprises two chromosomes and two plasmids. All protein coding regions in the OV14 genome were functionally grouped based on an eggNOG database. No genes homologous to the A. tumefaciens Ti plasmid vir genes appeared to be present in the OV14 genome. Unexpectedly, OV14 and 1021 were found to possess homologs to chromosomal based genes cited as essential to A. tumefaciens T-DNA transfer. Of significance, genes that are non-essential but exert a positive influence on virulence and the ability to genetically transform host genomes were identified in OV14 but were absent from the 1021 genome. Conclusions This study reveals the presence of homologs to chromosomally based Agrobacterium genes that support T-DNA transfer within the genome of OV14 and other alphaproteobacteria. The sequencing and analysis of the OV14 genome increases our understanding of T-DNA transfer by non-Agrobacterium species and creates a platform for the continued improvement of Ensifer-mediated transformation (EMT).
    • Insights into the transcriptomic response of the plant engineering bacterium Ensifer adhaerens OV14 during transformation

      Zuniga-Soto, Evelyn; Doohan, Fiona; Mullins, Ewen; Fitzpatrick, David; Science Foundation Ireland; Teagasc Walsh Fellowship Programme; 11/RFP.1/GEN/3420; 2011210 (Springer Science and Business Media LLC, 2019-07-17)
      The ability to engineer plant genomes has been primarily driven by the soil bacterium Agrobacterium tumefaciens but recently the potential of alternative rhizobia such as Rhizobium etli and Ensifer adhaerens OV14, the latter of which supports Ensifer Mediated Transformation (EMT) has been reported. Surprisingly, a knowledge deficit exists in regards to understanding the whole genome processes underway in plant transforming bacteria, irrespective of the species. To begin to address the issue, we undertook a temporal RNAseq-based profiling study of E. adhaerens OV14 in the presence/absence of Arabidopsis thaliana tissues. Following co-cultivation with root tissues, 2333 differentially expressed genes (DEGs) were noted. Meta-analysis of the RNAseq data sets identified a clear shift from plasmid-derived gene expression to chromosomal-based transcription within the early stages of bacterium-plant co-cultivation. During this time, the number of differentially expressed prokaryotic genes increased steadily out to 7 days co-cultivation, a time at which optimum rates of transformation were observed. Gene ontology evaluations indicated a role for both chromosomal and plasmid-based gene families linked specifically with quorum sensing, flagellin production and biofilm formation in the process of EMT. Transcriptional evaluation of vir genes, housed on the pCAMBIA 5105 plasmid in E. adhaerens OV14 confirmed the ability of E. adhaerens OV14 to perceive and activate its transcriptome in response to the presence of 200 µM of acetosyringone. Significantly, this is the first study to characterise the whole transcriptomic response of a plant engineering bacterium in the presence of plant tissues and provides a novel insight into prokaryotic genetic processes that support T-DNA transfer.