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Series GSE1695 Query DataSets for GSE1695
Status Public on Aug 24, 2004
Title Synechocystis sp. PCC 6803 response to inorganic carbon limitation
Organism Synechocystis sp. PCC 6803
Experiment type Expression profiling by array
Summary Synechocystis 6803 cells was grown photoautotrophically at 32 °C buffered in BG-11 and bubbled with 3% CO2. A relatively mild Ci stress was applied by switching the aeration from 3% CO2 to air alone. After incubation under designated conditions, a 100-ml aliquot of culture was immediately combined with an equal volume of ice-cold mixture of phenol and ethanol (1:10, w/v) in an ice bath. The resultant cells were collected by centrifugation at 1000 x g for 10 min at 4 °C. Total RNA was isolated with RNeasy Midi Kit (Qiagen, Valencia, CA) and further treated with the DNA-free kit (Ambion, Austin, TX). Fluorescently labeled cDNA was produced via a two-step indirect procedure involving cDNA synthesis from 16 µg of total RNA in a reverse transcriptase reaction incorporating aminoallyl-modified deoxynucleotide, followed by the second step involving chemical coupling of fluorescent dye to the introduced amino moieties of the synthesized cDNA. Labeled cDNA were adjusted to 14.75 µl, and the remainder of the hybridization components containing 2.5 µl of 10 µg µl-1 salmon sperm DNA, 8.75 µl of 20x SSC, 0.25 µl of 10% SDS, and 8.75 µl of formamide were added. The mixture was then heated for 2 min at 99 °C and maintained at 42 °C until hybridization. Hybridizations were preformed in a static incubator at 42 °C for 12-16 h then washed by placing in a 250-ml solution of 2x SSC and 0.1% SDS at 42 °C for 5 min with gentle agitation provided by rotation of a magnetic stir bar. The slide was transferred quickly to a 250-ml solution of 0.1x SSC and 0.1% SDS, incubated for 10 min at room temperature with gentle agitation, and washed five additional times in 0.1x SSC for 1 min at room temperature. Hybridization signals from the microarray were quantified using GenePix Pro 4.1 (Axon Instruments, Union City, CA). The quality control procedures were conducted in the image analysis software, and then data were saved to Acuity 3.1 (Axon Instruments).
Keywords: time-course
 
 
Contributor(s) Wang H, Postier BL, Burnap RL
Citation(s) 14612435
Submission date Aug 21, 2004
Last update date Mar 15, 2012
Contact name Robert Burnap
E-mail burnap@biochem.okstate.edu
Phone 405-744-7445
Organization name Oklahoma State University
Street address
City Stillwater
State/province OK
ZIP/Postal code 74078
Country USA
 
Platforms (1)
GPL965 Synechocysis sp. PCC6803 full genome array version 1
Samples (16)
GSM29242 60min_Rep1
GSM29243 60min_Rep2
GSM29244 60min_Rep3
Relations
BioProject PRJNA90659

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