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Sample GSM976940 Query DataSets for GSM976940
Status Public on Dec 06, 2012
Title NgnrGR plusDex plusGem, rep2
Sample type RNA
Source name Ectoderm co-injected with RNAs encoding NgnrGR and geminin, with Ngnr1 activity induced by Dex addition
Organism Xenopus laevis
Characteristics tissue: embryonic ectoderm
rna injection: Dex-inducible version of Ngnr1 (NgnrGR) and geminin
treatment: dexamethasone
Treatment protocol When sibling embryos were at stages 10–10.5, explants were pretreated with cycloheximide (final concentration 10mg/ml) for 30min and dexamethasone (final 10 mM) was added to induce transcriptional activity of Ngnr-GR as described previously (EMBO J.26(24):5093-5108). Explants were incubated at 25 degrees C for 2.5 hours and frozen in liquid nitrogen (when sibling embryos were at stage 12.0–12.5).
Growth protocol Both blastomeres of two-cell-stage pigmented embryos were injected with X-Ngnr-GR (10pg) with or without geminin (100pg) RNAs (with embryos injected with beta-galactosidase RNA as controls) and were raised until stages 8–9. Xenopus animal caps (50–60 ectodermal explants) per sample were isolated and raised in 0.7MMR at 25 degrees C.
Extracted molecule total RNA
Extraction protocol Total RNA was prepared with Trizol (Invitrogen) (Washington University Genome Technology Access Center).
Label biotin
Label protocol Total RNA (20mg) per sample was used for probe synthesis and hybridization to Affymetrix Xenopus laevis Genome Arrays (Washington University Genome Technology Access Center).
Hybridization protocol Performed by the Washington University Genome Technololgy Access Center.
Scan protocol Affymetrix GeneChip 3000 used by the Washington University Genome Technololgy Access Center.
Data processing Result was analyzed with dChip software ( Raw CEL and DAT files were analyzed with dChip software ( after normalization. The entire experiment (microinjection, RNA extraction, and microarray analysis) was repeated two times and quantitative RT-PCR was used to determine cut-offs (see below). Target genes were determined by comparing microarray and quantitative real-time RT-PCR (qRT-PCR) data. Candidates were preliminarily determined from microarray data based on the following criteria: i) expression, represented by model-based expression indices (MBEI), was increased more than 1.2-fold in DEX-treated groups (E/B > 1.2); ii) MBEI differences were larger than 50 (E-B > 50); and iii) expression was claimed as 'present (P) in at least one sample in each experiment. Genes identified as regulated in both experiments were considered candidates. Genes induced in DEX-treated, beta-galactosidase-injected samples compared with the DEX-untreated, Ngnr1-GR-injected samples were also determined and excluded from analysis as they were regarded as DEX-induced. From Ngnr1 candidate target lists, we used qRT-PCR to determine fold-change cut-off values. We found that candidate genes showing E/B > 1.5 and E-B > 70 for Ngnr1 or E/B > 1.3 consistently showed more than 2-fold induction by qRT-PCR.
Submission date Jul 25, 2012
Last update date Jan 23, 2014
Contact name Kristen L Kroll
Organization name Washington University School of Medicine
Department Developmental Biology
Lab Kristen Kroll
Street address 320 McDonnell Sciences/660 S. Euclid Ave.
City Saint Louis
State/province MO
ZIP/Postal code 63110
Country USA
Platform ID GPL1318
Series (2)
GSE39658 Regulation of neurogenin-dependent gene expression by geminin
GSE39673 Geminin regulates the transcriptional and epigenetic status of neuronal fate promoting genes during mammalian neurogenesis

Data table header descriptions
VALUE dChIP-normalized signal intensity

Data table
Xl.326.1.S1_at 9.96
Xl.17432.1.S1_at 4177.82
Xl.17432.1.S2_at 222.73
Xl.7551.2.S1_a_at 1098.33
Xl.7551.2.S1_x_at 1107.76
Xl.7551.1.S1_x_at 887.26
Xl.8672.1.S1_at 337.99
Xl.8805.1.S1_at 230.36
Xl.8805.1.S1_s_at 1082.9
Xl.11405.1.S1_a_at 13.43
Xl.4515.1.S1_at 3479.26
Xl.4515.1.S2_a_at 865.97
Xl.2168.1.S1_s_at 1068.69
Xl.1728.1.S1_at 1610.56
Xl.4111.1.S1_at 2599.75
Xl.23295.1.S1_a_at 2310.11
Xl.4150.1.S1_at 2543.94
Xl.3421.1.S1_at 4463.09
Xl.26147.1.A1_at 3381.13
Xl.26147.1.A1_s_at 6451

Total number of rows: 15491

Table truncated, full table size 344 Kbytes.

Supplementary file Size Download File type/resource
GSM976940_xNgnr1-GR_plusDex_plusGem_rep2.CEL.gz 2.0 Mb (ftp)(http) CEL
Processed data included within Sample table

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