Halobacterium NRC-1 Growth curve and GTF perturbation
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ABSTRACT:
ORGANISM(S): Halobacterium salinarum NRC-1
PROVIDER: GSE6776 | GEO | 2007/02/23
SECONDARY ACCESSION(S): PRJNA98919
REPOSITORIES: GEO
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Project description:Halobacterium salinarum NRC-1 was grown in CM media, at 37oC in a waterbath with agitation of 125 rpm under constant light. Analysis of transcriptional changes during growth, in addition to mapping of transcriptome structure under the same conditions, provided interesting insights about regulatory logic within prokaryotic coding regions.
2009-06-25 | GSE12977 | GEO
Project description:Halobacterium cells, specified general transcription factor knock-out strains or plasmid expressed 6-His tagged general transcription factor strains, were grown to mid-log phase. RNA extractions were performed using the Stratagene Absolutely RNA Miniprep Kit and RNA quality checked with the Agilent Bioanalyzer and with Oligo Microarrays were fabricated at the Institute for Systems Biology Microarray Facility. The arrays contain 4 spots per unique 70-mer oligonucleotides for each of 2400 non-redundant genes in Halobacterium NRC-1 Labeling, hybridization and washing have been previously described (Baliga et al. 2002) with 10 μg of RNA from the sample and reference. RNA from the final time point of a replicate experiment was used as the reference. . Bias in dye incorporation was accounted for by reversing the labeling dyes (dye-flip). Raw data was processed and converted into log10 ratios with lambda (λ) values determined by a maximum likelihood method. Baliga, N. S., Pan, M., Goo, Y. A., Yi, E. C., Goodlett, D. R., Dimitrov, K., Shannon, P., Aebersold, R., Ng, W. V. & Hood, L. (2002) Proc Natl Acad Sci U S A 99:14913-84. Keywords: genetic perturbation
2008-01-03 | GSE7713 | GEO
Project description:Halobacterium salinarum NRC-1 growth curve
| PRJNA114245 | ENA
Project description:Halobacterium salinarum NRC-1 growth curve, tiling arrays
| PRJNA114261 | ENA
Project description:Halobacterium NRC-1 Growth curve and GTF perturbation
| PRJNA98919 | ENA
Project description:Halobacterium cells, specified general transcription factor knock-out strains or plasmid expressed 6-His tagged general transcription factor strains, were grown to mid-log phase. RNA extractions were performed using the Stratagene Absolutely RNA Miniprep Kit and RNA quality checked with the Agilent Bioanalyzer and with Oligo Microarrays were fabricated at the Institute for Systems Biology Microarray Facility. The arrays contain 4 spots per unique 70-mer oligonucleotides for each of 2400 non-redundant genes in Halobacterium NRC-1 Labeling, hybridization and washing have been previously described (Baliga et al. 2002) with 10 μg of RNA from the sample and reference. RNA from the final time point of a replicate experiment was used as the reference. . Bias in dye incorporation was accounted for by reversing the labeling dyes (dye-flip). Raw data was processed and converted into log10 ratios with lambda (λ) values determined by a maximum likelihood method. Baliga, N. S., Pan, M., Goo, Y. A., Yi, E. C., Goodlett, D. R., Dimitrov, K., Shannon, P., Aebersold, R., Ng, W. V. & Hood, L. (2002) Proc Natl Acad Sci U S A 99:14913-84. each knockout or perturbed expression strain was analyzed at a single point during growth. This sample was analyzed on duplicate microarrays as a dye flip (2 microarrays per sample).
2010-05-27 | E-GEOD-7713 | biostudies-arrayexpress
Project description:Halobacterium NRC-1 General transcription factor perturbation
| PRJNA99845 | ENA
Project description:Halobacterium salinarum NRC-1 was grown in CM media, at 37oC in a waterbath with agitation of 125 rpm under constant light. Analysis of transcriptional changes during growth, in addition to mapping of transcriptome structure under the same conditions, provided interesting insights about regulatory logic within prokaryotic coding regions. Samples were collected at different cell densities, from OD ~0.2 to OD ~5.0. 2 biological replicates were conducted. For each sample, a dye-swap experiment was performed.
2010-05-17 | E-GEOD-12977 | biostudies-arrayexpress
Project description:Experimentally mapped transcriptome structure of H. salinarum NRC-1 by hybridizing total RNA (including RNA species <200 nt) to genome-wide high-density tiling arrays (60 mer probes with 40 nt overlap between contiguous probes). H. salinarum NRC-1 presents a number of interesting switches in metabolism during growth due to complex changes in EFs including pH, oxygen, nutrition, etc. While most single perturbations (radiation, oxygen, metals, etc.) affect the expression of only ~10% of all genes (Baliga et al, 2004; Kauret al , 2006; Whitehead et al, 2006), the changes during growth resulted in differential regulation of a significantly higher proportion of genes (~63%, 1,518 genes). These conditions enabled the investigation of a wider transcriptional landscape, which includes not only modulation of transcript levels, but also extensive changes in transcriptome structure. We observed altered transcription start sites (TSSs), transcription termination site (TTSs), operon organizations and differential regulation of putative ncRNAs. By integrating hybridization signals with dynamic growth-related changes, we estimated the probability that each tiling array probe was complementary to a transcribed region, mapped locations of putative transcript boundaries and identified 1,574 TSSs and 1,952 TTSs for most genes with some transcriptional variation. In sum, TSSs were assigned to 64% (1,156 singletons and 544 genes in 203 operons) of all annotated genes and TTSs were assigned to 1,114 genes and 202 operons. We were also able to identify 5' and 3' UTRs and revise start sites for 61 genes and 12 operons. H. salinarum NRC-1 growth curve experiments were conducted in CM media, in a water bath incubator at 37oC with agitation of 125 rpm. Reference samples were cultured under standard growth conditions (Baliga and DasSarma, 1999), at mid-log phase (OD600 = ~0.6), as well as all strains used for ChIP-chip experiments (Facciotti et al., 2007). Whole-genome high resolution tiling arrays for H. salinarum NRC-1 were designed with e-Array (Agilent Technologies), using strand specific 60mer probes tiled every 20nt for the main chromosome (NC_002607) and every 21nt for the plasmids pNRC200 (NC_002608) and pNRC100 (NC_001869), consisting a total of 244K probes, including manufacturersâ?? controls. Microarrays were printed by Agilent technologies and hybridized to total RNA, which was isolated using mirVana miRNA Isolation kit (Ambion) and direct labeled with Alexa547 and Alexa647 dyes (Kreatech) (Baliga et al., 2004). We used direct chemical labeling of RNA (Baliga et al., 2004) to avoid enzymatic labeling artifacts (Perocchi et al., 2007) and enable strand-specific signals for transcribed segments. Hybridization and washing were performed according to array manufacturerâ??s instructions. Arrays were scanned in ScanArray (Perkin Elmer) and spot-finding was performed using Feature Extraction (Agilent Technologies). Two biological replicates were sampled and dye-flip experiments were conducted for each sample. Resulting intensities were quantile normalized across all experiments. Log ratios were calculated for each probe (growth curve sample/reference). Reference RNA signals were normalized by sequence content using a linear model similar to that of (Johnson et al., 2006). Interactive visualization of the data was performed in the Gaggle Genome Browser (Bare et al., in preparation), available at http://baliga.systemsbiology.net/regulatory_logic/.
2010-05-17 | E-GEOD-12923 | biostudies-arrayexpress
Project description:ChIP-Chip of General Transcription factors in Halobacterium NRC-1
| PRJNA114681 | ENA