Halobacterium NRC-1 Diurnal experiment Dark 3
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ABSTRACT: The wild type strain of Halobacterium salinarium strain NRC-1 was used for the diel growth experiments. Culturing from colony was done under constant light in a liquid Complete Medium (CM; at 37 ºC with shaking at 220 rpm). For experiment, cultures were started at an OD = 0.005. Cultures were grown aerobically (shaking at 120 rpm) and a temperature of 37° C under light:dark cycles (12:12 hrs) for 72 hours then grown in complete darkness. Light intensity was 150 μE/m2/sec. Time course sampling began after the 72 hour light:dark phase. A second culture was also sampled as a control and was grown under complete darkness (no light:dark cycle) before sampling began. Samples from both the experimental and control cultures were collected after 83 hours of exposure to the light:dark cycle every 1 to 3 hours for another 25 hours. Aliquots of the cultures were centrifuged and the cell pellet flash-frozen in liquid nitrogen after decanting the supernatant. RNA extractions were performed using the Stratagene Absolute RNA kit and RNA quality checked with the Agilent Bioanalyzer and with PCR. 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 5 μg of RNA from the sample and 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. 24 conditions were analyzed on duplicate microarrays as dye-flips (48 total microarrays).
ORGANISM(S): Halobacterium sp. NRC-1
SUBMITTER: Kenia Whitehead
PROVIDER: E-GEOD-7711 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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