Comment[ArrayExpressAccession]	E-GEOD-42018
MAGE-TAB Version	1.1							
Public Release Date	2012-11-07
Investigation Title	Cerebellar Granule Neuron Temporal-NFI Microarray Data							
Comment[Submitted Name]	Cerebellar Granule Neuron Temporal-NFI Microarray Data
Experiment Description	Dendrite and synapse development are critical for establishing appropriate neuronal circuits, and disrupted timing of these events can alter connectivity leading to disordered neural function. In early postnatal development of cerebellar granule neurons (CGNs), the expression of many genes is temporally regulated. Further, NFI (Nuclear Factor I) proteins have been shown to play important roles in the regulation of gene expression in developing CGNs. To identify NFI-regulated genome-wide targets involved in late maturation of mouse CGNs, we performed two groups of microarray expression analysis: (1) temporal expression arrays using 1.5 and 6 day cultures of mouse CGNs representing immature and more mature CGNs, respectively, which identified 844 Temporal-Up or Down genes; and (2) NFI-regulated genes in 6 day CGN cultures that were infected at the time of plating with lentiviral vectors expressing either NFI dominant repressor (NFI-EnR) or EnR control protein. This identified 686 NFI-Up (NFI-EnR down-regulated) or NFI-Down (NFI-EnR up-regulated) genes. Overlap analysis identified 212 temporal genes that were regulated by NFI. These results indicated that NFI plays a pivotal role in the regulation of late CGN maturation. For temporal arrays,  mouse CGN progenitors were purified from P6 mouse cerebellum and cultured for either 1.5 or 6 days. Three biological replicates were analyzed for each time point. For NFI arrays, CGN progenitors from P6 mice were transduced upon plating with either NFI-EnR or EnR control lentivirus and cultured 6 days. Four pairs of biological replicates were performed.							
Term Source Name	ArrayExpress	EFO
Term Source File	http://www.ebi.ac.uk/arrayexpress/	http://www.ebi.ac.uk/efo/efo.owl						
Person Last Name	Ding	Ding	Wang	Xi	Kilpatrick			
Person First Name	Baojin	Baojin	Wei	Hualin	Daniel			
Person Mid Initials				S	L			
Person Email	Baojin.Ding@umassmed.edu							
Person Affiliation	UMASS Medical School							
Person Phone	508-856-2679							
Person Address	Microbiology and Physiological Systems , UMASS Medical School, 55 Lake AVE. N., Worcester, MA, USA							
Person Roles	submitter							
Protocol Name	P-GSE42018-1	P-GSE42018-6	P-GSE42018-3	P-GSE42018-8	P-GSE42018-7	P-GSE42018-2	P-GSE42018-4	P-GSE42018-5
Protocol Description	ID_REF =  VALUE = Quantile normalized gene level expression values from RMA.	Samples were hybridized usingM- the Affymetrix Hybridization Cocktail recipe consisting of components (BSA 50mg/ml from Invitrogen and Herring Sperm DNA from Promega) and lab prepared 2X Hybridization Buffer M-bM-^@M-" Pre-hybridize arrays at 450C for 10 minutes with 1X Hybridization Buffer Heat cocktails at 99M-0 for 5 minutes, then 45M-0 for 5 minutes and centrifuge at max speed for ~3 minute (N.B. this deviates fromM- Affy SOP)M-bM-^@M-" TransferM- 200 M-NM-<l ofM- hybridization solution to each array, then cover holes with Tough Spots M-bM-^@M-" Hybridize ~16 hours at 45M-0C at 60rpmM-bM-^@M-" Fluidics wash protocol used was EukGE-WS2v5.	Mouse CGN progenitors were purified from P6 mice cerebellum and cultured in complete Neurobasal medium until 1.5 or 6 days.	NFI-EnR data were processed using RMA. Gene expression values in the Affymetrix cel files were first background corrected and normalized together using RMA algorithm (Bioconductor R package). Low expression probes (i.e., expression value below 40th percentile in at least 7 out of 8 arrays) were removed. For multiple probes mapping to the same gene, only the probe showing the highest expression change for each gene was retained. Finally, moderated t-statistics and FDR correction (limma R package v.2.15.1.) were used to assess the significance of differential expression. Identical steps for signal normalization, background noise correction and statistical testing were applied to temporal arrays. NFI differentially regulated genes were defined as probes with FDR P-value<0.15 and fold change>1.5. Differentially expressed temporal genes were defined as having probes with FDR P-value<0.1 and fold change>2.	Affymetrix Gene ChIP Scanner 3000 7G	For NFI group, CGNs were transduced on the day of plating with lentiviruses expressing either NFI-EnR or EnR alone. Fresh medium was added on 3DIV (days in vitro).	RNA was extracted from cultured cells using Tri reagent (Sigma) and purified with an RNeasy Plus Micro Kit (QIAGEN).	Samples were prepared using an Affymetrix GeneChip 3M-bM-^@M-^YIVT Express Kit, in which total RNA undergoes reverse transcription to synthesize first strand cDNA and is then converted into double-stranded DNA template for transcription. IVT synthesizes aRNA and incorporates a biotin-conjugated nucleotide. The aRNA is then purified and fragmented.
Protocol Type	bioassay_data_transformation	hybridization	grow	feature_extraction	image_aquisition	specified_biomaterial_action	nucleic_acid_extraction	labeling
Experimental Factor Name	TRANDUCED WITH	DAY IN VITRO DIV						
Experimental Factor Type	tranduced with	day in vitro div						
Comment[SecondaryAccession]	GSE42018							
Comment[GEOReleaseDate]	2012-11-07							
Comment[ArrayExpressSubmissionDate]	2012-11-02							
Comment[GEOLastUpdateDate]	2012-11-08							
Comment[AEExperimentType]	transcription profiling by array							
Comment[AdditionalFile:Data1]	GSE42018_differentially_expressed_genes.xlsx							
SDRF File	E-GEOD-42018.sdrf.txt