biostudies-arrayexpressMetabolomicsUnknownTranscriptomicsGenomicsProteomicsChris Seideltranscription profiling by arraySchmidtea mediterraneaSchmidtea mediterraneahttps://www.ebi.ac.uk/biostudies/studies/E-GEOD-56181Planarian flatworms can regenerate every organ after amputation. Adult pluripotent stem cells drive the ability to regenerate, but how injury activates these cells and directs them into the appropriate lineages is not understood. To study the regeneration response in a simplified context, we selectively induced the ejection of the planarian pharynx by briefly exposing animals to sodium azide followed by microarray analysis. Tissue surrounding the wound site was examined at 6, 12, 18, 24, 48, and 72 hours post amputation by comparison to tissue isolated immediately after amputation (time zero). Stem cells are required for the regeneration process, and can be eradicated by exposure to radiation. A parallel time course was also performed on worms that had been irradiated with 10k rads of gamma irradiation. Two time courses consisting of irradiated or unirradiated worms following chemical amputation of the pharynx. Tissue samples taken at 6, 12, 18, 24, 48 and 72 hours are each compared to a reference sample at time zero. The experiment was performed in triplicate for a total of 42 samples on 36 arrays.biostudies-arrayexpressGrowth Protocol - Schmidtea mediterranea asexual clonal line CIW4 was maintained and used as previously described (Newmark and Sánchez Alvarado, 2000).Nucleic Acid Extraction - At specified times after amputation, plugs were extracted using 1mm microcapillary pipets (FHC, catalog # 30-30-0), and transferred directly into Trizol (Life Technologies) using a mouth pipet. For each replicate, 25 plugs were homogenized together, and then chloroform-extracted. The pellet was then precipitated with isopropanol, washed, and resuspended in water. RNA was then passed over an RNEasy column and DNasetreated.Scaning - Arrays were scanned using an Agilent G2505C scanner and quantified with Agilent Feature Extraction software version 10.5 using protocol GE2_105_Dec08.Labeling - Starting with 100 ng total RNA, amplification and labeling with Cy3 or Cy5 was performed using the Low Input Quick Amp Labeling Kit Two-Color from Agilent Technologies (#5190-2306).Hybridization - Custom Agilent 4x44k arrays with design id: 033226 were hybridized using oligoarray control targets and Agilent hybridization buffers according to the manufacturer protocols.Sample Treatment - Irradiated animals were exposed to 10,000 rads on a GammaCell 40 Exactor irradiator. Planarian water was replaced with 100mM sodium azide (diluted in Montjuïc water). After 5-7 minutes, the pharynx was visibly extended out of the body. Vigorous pipetting often dislodged the pharynx from the body; if necessary, fine serrated forceps were used to remove the pharynx, followed by several washes in Montjuïc.MIAME ScoreRaw DataOrganizationAssays and DataProcessed DataMAGE-TAB FilesArray DesignsA Sanchez AlvaradoChris SeidelSean McKinneyCarolyn AdlerData Transformation - Data was analyzed in the R environment using the Limma library (Smyth, 2004) for loess normalization and calculation of p-values between treatments. ID_REF = VALUE = Normalized log10 ratio (Cy5/Cy3) representing each timepoint/time zero.falseSelective amputation of the pharynx identifies a FoxA-dependent regeneration program in planariaPlanarian flatworms can regenerate every organ after amputation. Adult pluripotent stem cells drive the ability to regenerate, but how injury activates these cells and directs them into the appropriate lineages is not understood. To study the regeneration response in a simplified context, we selectively induced the ejection of the planarian pharynx by briefly exposing animals to sodium azide followed by microarray analysis. Tissue surrounding the wound site was examined at 6, 12, 18, 24, 48, and 72 hours post amputation by comparison to tissue isolated immediately after amputation (time zero). Stem cells are required for the regeneration process, and can be eradicated by exposure to radiation. A parallel time course was also performed on worms that had been irradiated with 10k rads of gamma irradiation. Two time courses consisting of irradiated or unirradiated worms following chemical amputation of the pharynx. Tissue samples taken at 6, 12, 18, 24, 48 and 72 hours are each compared to a reference sample at time zero. The experiment was performed in triplicate for a total of 42 samples on 36 arrays.2014-03-26T00:00:00Z2023-09-08T03:55:12.105Z2022-02-02T22:51:16.763ZE-GEOD-56181GSE56181EFO_0002768