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Sample GSM976939 Query DataSets for GSM976939
Status Public on Dec 06, 2012
Title NgnrGR plusDex, rep2
Sample type RNA
Source name Ectoderm injected with RNA encoding NgnrGR, and treated with Dex to induce Ngnr-dependent target gene induction
Organism Xenopus laevis
Characteristics tissue: embryonic ectoderm
rna injection: Dex-inducible version of Ngnr1 (NgnrGR)
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 11.91
Xl.17432.1.S1_at 5262.95
Xl.17432.1.S2_at 156.32
Xl.7551.2.S1_a_at 1459.13
Xl.7551.2.S1_x_at 1426.11
Xl.7551.1.S1_x_at 1185.37
Xl.8672.1.S1_at 430.81
Xl.8805.1.S1_at 272.53
Xl.8805.1.S1_s_at 1475.26
Xl.11405.1.S1_a_at 12.11
Xl.4515.1.S1_at 4104.34
Xl.4515.1.S2_a_at 158.26
Xl.2168.1.S1_s_at 2133.59
Xl.1728.1.S1_at 1887.07
Xl.4111.1.S1_at 4746.44
Xl.23295.1.S1_a_at 2660.07
Xl.4150.1.S1_at 2391.18
Xl.3421.1.S1_at 4153.31
Xl.26147.1.A1_at 3131.28
Xl.26147.1.A1_s_at 6330.65

Total number of rows: 15491

Table truncated, full table size 344 Kbytes.

Supplementary file Size Download File type/resource
GSM976939_xNgnr1-GR_plusDex_rep2.CEL.gz 2.0 Mb (ftp)(http) CEL
Processed data included within Sample table

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