Selected target genes for expression analysis by qRT-PCR.

<p>Selected target genes for expression analysis by qRT-PCR.</p

Effect of AbsR25 sRNA overexpression.

<p>Comparison of expression of predicted putative target genes in <i>A. baumannii</i> cells harbouring recombinant plasmid over-expressing AbsR25 sRNA. Expression of target genes was studied under arabinose induced and uninduced conditions. The experiments were performed in triplicates.</p

List of putative 31 sRNAs and their features.

<p>Up ORF: Upstream Open Reading frame; dist Up ORF: distance of sRNA from Upstream Open Reading frame; UpORF dir: Upstream Open Reading frame direction; dir sRNA: direction of small RNA expression; len sRNA: length of sRNA; dist Dn ORF: distance of sRNA from downstream open reading frame.</p

Northern blot showing expression of AbsR25 small RNA under different stress conditions.

<p>Northern blot showing the expression of AbsR25 small RNA candidate in Nutrient broth medium at different stress conditions is presented. (A) Total RNA was extracted from <i>A. baumannii</i> MTCC1425 cells growing at different temperature conditions (30°C, 37°C, 45°C, 50°C). 25 μg of total RNA sample of temperature stresses; 30°C, 37°C, 45°C, 50°C were loaded in lane 1 to 4 respectively. (B) Total RNA was extracted from <i>A. baumannii</i> MTCC1425 cells growing in nutrient broths containing 100, 200, 300, 400 and 500 mM of NaCl. 25 μg of extracted total RNA from above mentioned concentrations of NaCl were loaded from lane 1 to 5 respectively. (C) 25 μg of extracted total RNA of <i>A. baumannii</i> MTCC1425 cells growing in 0 μg/ml, 64 μg/ml, 128 μg/ml, 192 μg/ml and 256 μg/ml supplemented concentration of EtBr were loaded on PAGE from lane 1 to 5 respectively. Riborular low range RNA ladder (Fermentas, USA) sizes are shown to the left of the blot. Small RNA was detected by biotin labeled riboprobes designed to pair with the predicted small RNA. Corresponding ethidium bromide stained 5S rRNA was used as loading control to ensure equal loading.</p

Real time expression of selected target genes.

<p>Relative expression of <i>in silico</i> predicted putative target genes [A1S_3401 (hypothetical protein), A1S_3251 (amino acid transporter LysE), A1S_1791 (putative tartrate symporter MFS superfamily protein), A1S_2269 (RtcR - Regulator of RNA terminal phosphate cyclase), A1S_1505 (hypothetical protein), A1S_0229 (hypothetical protein), A1S_3342 (putative arsenate reductase), A1S_1627 (hypothetical protein), A1S_2584 (major facilitator superfamily family drug transporter), A1S_2660 (RND efflux transporter), A1S_1331 (major facilitator superfamily transporter)] of AbsR25 sRNA under different concentrations of EtBr.</p

Distribution and conservation of the identified sRNA gene sequence (<i>absR25</i>) in closely related bacteria.

<p>Here in table; Aba, Acinetobacter baumannii; Aca, Acinetobacter calcoaceticus; Aol, Acinetobacter oleivorans; N, no similar sequence found.</p

Selected target genes for expression analysis by qRT-PCR.

<p>Selected target genes for expression analysis by qRT-PCR.</p

Oligonucleotides used in this study.

<p>Oligonucleotides used in this study.</p

The expression of novel <i>A. baumannii</i> small RNA in different growth phases.

<p>Northern blot was used for determining the intracellular abundance of AbsRs over the course of growth (lag phase, L; early exponential phase, EE; exponential phase, E; stationary phase, S). Riborular low range RNA ladder (Fermentas, USA) sizes are shown to the left of the blot. Corresponding ethidium bromide stained 5S rRNA was used as loading control to ensure equal loading and shown below each Northern blot. (A) Northern blot of AbsR11, (B) Northern blot of AbsR25, (C) Northern blot of AbsR28.</p

Schematic representation of bioinformatic approach used for small RNA species prediction in <i>Acinetobacter baumannii</i> ATCC 17978.

<p>The first step was to determine the inter-genic regions (IGRs) from the annotated ORFs (Open Reading frames) and then running them through BLAST against genomes of related species. The conserved sequences were subjected to QRNA program wherein conserved secondary structure regions among the pairwise aligned sequences were determined.</p