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Sample GSM976935 Query DataSets for GSM976935
Status Public on Dec 06, 2012
Title NgnrGR, rep 1
Sample type RNA
Source name Xenopus ectoderm injected with RNA encoding NgnrGR
Organism Xenopus laevis
Characteristics tissue: embryonic ectoderm
rna injection: Dex-inducible version of Ngnr1 (NgnrGR)
treatment: no 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 10.52
Xl.17432.1.S1_at 3376.93
Xl.17432.1.S2_at 154.01
Xl.7551.2.S1_a_at 929.15
Xl.7551.2.S1_x_at 815.82
Xl.7551.1.S1_x_at 865.71
Xl.8672.1.S1_at 310.36
Xl.8805.1.S1_at 101.89
Xl.8805.1.S1_s_at 1372.44
Xl.11405.1.S1_a_at 5.9
Xl.4515.1.S1_at 2755.24
Xl.4515.1.S2_a_at 95.27
Xl.2168.1.S1_s_at 1681.16
Xl.1728.1.S1_at 1423.74
Xl.4111.1.S1_at 2630.66
Xl.23295.1.S1_a_at 2314.27
Xl.4150.1.S1_at 1145.75
Xl.3421.1.S1_at 2986.23
Xl.26147.1.A1_at 2746.17
Xl.26147.1.A1_s_at 4267.89

Total number of rows: 15491

Table truncated, full table size 342 Kbytes.

Supplementary file Size Download File type/resource
GSM976935_NgnrGR_rep1.CEL.gz 1.9 Mb (ftp)(http) CEL
Processed data included within Sample table

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